Nematicidal effect of rhizobacteria on plant-parasitic nematodes associated with vineyards.

The action of metabolites and exoenzymes from rhizobacteria on different plant-parasitic nematodes has an influence on the nematicidal efficacy of the microbe. Seven rhizobacteria, divided into two bacterial groups, were evaluated in vitro for nematicidal activity on Meloidogyne ethiopica and Xiphinema index. The direct effect of their filtrates on egg hatching and juveniles of M. ethiopica as well as mobile stages of X. index was evaluated during a 72-h period. The production of four exoenzymes and two metabolites associated with nematode mortality was investigated. Molecular characterization of three isolates was performed, and the physiological profiles and lipase activity of all isolates were obtained using the BIOLOG EcoPlate system. While chitinase and collagenase were measured using the BIOLOG MT2 plate system, protease, hydrogen cyanide and hydrogen sulphide were directly determined in Petri dishes. Nematode mobile stages exposure to the bacterial filtrate revealed a nematicidal effect up to 93.7% on X. Index and up to 83.3% on M. ethiopica. The control of egg hatching varied between 35 and 85%. A positive correlation was found between the mortality of both nematode mobile stages and the concerted activities of the bacterial enzymes as well as the level of the volatile metabolites. The nematicidal effect of rhizobacteria strains varies by nematode genera and among the developmental stages evaluated.

Neonatal Serial Transverse Enteroplasty (STEP): Case Report.

CASE REPORT: Gastroschisis is the most frequent congenital abdominal wall defect. When associated with intestinal atresia (complex gastroschisis), short bowel syndrome may occur. Complicated gastroschisis is the most frequent cause of short bowel syndrome in our series. The serial transverse enteroplasty procedure has been used to lengthen the bowel and achieve intestinal rehabilitation in patients with dilated gut. The use of this technique in the newborn period, for tailoring the bowel while preserving absorptive mucosa, has been recently described. We present a video showing the surgical treatment of an intestinal obstruction produced by a complex intestinal atresia in a newborn baby in whom a primary closure of a gastroschisis had been done at birth. During laparotomy at the 21st day of life, a type IVa intestinal atresia was found, associated with a colonic stenosis. Proximal dilated jejunum was tailored with a serial transverse enteroplasty procedure, as shown in the video. End-to-end jejunal-ileal anastomosis was performed. Postoperative entero-cutaneous fistula occurred and was treated with vacuum-assisted therapy. Enteral feedings were initiated at 15 days after surgery. Parenteral nutrition was withdrawn at 30 days. After 16 months follow-up, actual weight was 8.7 kg (percentile 10% to 25%). The patient remained on full enteral feedings. CONCLUSIONS: In cases of intestinal atresia, short bowel syndrome, and proximal dilated bowel, we propose an aggressive surgical approach to preserve bowel mucosal surface while tailoring the jejunal loop to improve motility. The serial transverse enteroplasty procedure is an acceptable alternative to tailoring methods that resect a segment of the bowel wall.

Exoenzymes and metabolites related to the nematicidal effect of rhizobacteria on Xiphinema index Thorne & Allen.

AIMS: To identify enzymes and metabolites in the rhizobacteria filtrates that have a nematicidal effect on Xiphinema index and perform molecular characterization of the strains evaluated. METHODS AND RESULTS: A series of four bacteria selected for their nematicidal potential were considered for in vitro, biochemical and molecular studies. The direct effect of the bacterial filtrates was evaluated in vitro on X. index juveniles and adults. Hydrogen sulphide and hydrogen cyanide liberation and protease, chitinase, collagenase and lipase activity were verified in the strains. Up to five housekeeping genes and one ITS 16S-23S rRNA were analysed. All bacterial filtrates presented 54-100% mortality when evaluated during up to 72 h of nematode exposure. Strains presented protease activity; two of them (strains FB833T and FR203A) showed reliable collagenase and chitinase activities, respectively, and three of them showed strong lipolytic activity (FB833T, FR203A and FS213P). Strain Bacillus megaterium FB133M had no lipase activity and presented the lowest nematicidal effect. Bacillus amyloliquefaciens FR203A had the largest lethal effect. CONCLUSION: The rhizobacteria strains evaluated in this study possess nematicidal compounds, which may offer an interesting alternative for X. index control. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of exoenzymes and metabolites associated with nematicidal effect of rhizobacteria on X. index, which can be a possible alternative for control of this plant-parasitic nematode.

Phytoplasmas Associated with Grapevine Yellows Disease in Chile.

An extensive survey was performed from 2002 to 2006 to detect and identify phytoplasmas associated with Chilean grapevines. Nested polymerase chain reaction assays using phytoplasma universal primer pairs P1/P7 and R16F2n/R2 detected phytoplasmas in 34 out of the 94 samples tested (36%). Restriction fragment length polymorphism (RFLP) analyses, cloning, and sequencing allowed identification of phytoplasmas belonging to ribosomal subgroups 16SrI-B, 16SrI-C, 16SrVII-A, and 16SrXII-A. The 16SrVII-A phytoplasma represents a new finding in grapevine; moreover, variability of the RFLP profile was observed in some of the 16SrXII-A phytoplasmas, indicating possible new ribosomal subgroups. Mixed phytoplasma infections and infections of phytoplasmas together with one or more viruses also occurred.

Domain-restricted expression of two glutamic acid decarboxylase genes in midgestation mouse embryos.

Glutamic acid decarboxylase (GAD) is the biosynthetic enzyme for gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the central nervous system (CNS) of vertebrates. In addition to the adult CNS, GABA and GAD also have been detected in embryos, although their precise localization and specific functions in embryonic development have not been elucidated. In this paper, the authors studied the cellular distribution of two GAD isoforms, GAD65 and GAD67, in midgestation mouse embryos by in situ hybridization histochemistry. With few exceptions, it was found that GAD65 and GAD67 mRNAs are localized in overlapping cellular domains of the embryonic CNS that later develop into regions with a strong GABAergic contribution. The GAD-expressing cells are situated in the differentiating zone of the embryonic day 10.5 (E10.5) through E11.5 CNS and in the subventricular zone and the mantle zone of the E12.5 CNS, which suggests that they are committed neuronal precursors. By using a specific serum for GABA, a similar pattern of distribution was obtained, indicating that GAD mRNAs are translated efficiently into enzymatically active GAD, which produces embryonic GABA. The expression domains of GAD overlap with those of genes that are known to be involved in the patterning of the embryonic CNS. The two GAD mRNAs also are detected outside of the embryonic CNS in various cell types, mainly those of placodal and neural crest origin. This pattern of expression is consistent with the notion that GAD and its product, GABA, play a signaling role during development.

Biventricular repair for double-outlet right ventricle. Results and long-term follow-up.

BACKGROUND: The purpose of the present study was to define the optimal management and to identify the risk factors for death and repeat operation in patients with double-outlet right ventricle. METHODS AND RESULTS: From 1985 through 1996, 154 consecutive patients underwent biventricular repair for double-outlet right ventricle. The presence of bilateral infundibular structures was the major inclusion criteria (142 patients). According to the relationship of the ventricular septal defect (VSD) to the great arteries, there were 86 patients with a subaortic VSD (56%), 45 patients with a subpulmonary VSD (29%), 18 patients with a noncommitted VSD (12%), and 5 patients with a doubly committed VSD (3%). Sixty-five patients (42%) had undergone previous palliative procedures. At repair, the median age was 10 months, and the median weight was 6.5 kg. Two main types of repair were used: intraventricular baffle repair (n = 115) and arterial switch operation with VSD-to-pulmonary artery baffle (n = 39). There were 14 hospital deaths (9%; 70% confidence limit [CL], 7% to 12%). The only significant risk factor for early death was the presence of congenital mitral valve anomalies (P = 0.02). Twenty-eight patients (18%) required 39 repeat operations. The repeat operation rate was higher in patients with associated VSD enlargement at baffle construction (n = 29; 19%) (P = 0.01). There were 6 late deaths (4%; 70% CL, 2% to 7%). Patients presenting with pulmonary stenosis constituted a low-risk group for global death (P = 0.008). The median follow-up was 52 months. Ten-year actuarial survival and survival with freedom from repeat operation rates were 86% and 62% (70% CL, 83% to 89% and 54% to 70%), respectively. CONCLUSIONS: Long-term survival with good quality of life can be achieved after either 1- or 2-stage repair of this complex anomaly.

Beta-galactosidase-labelled relay neurons of homotopic olfactory bulb transplants establish proper afferent and efferent synaptic connections with host neurons.

The vertebrate olfactory system has long been an attractive model for studying neuronal regeneration and adaptive plasticity due to the continuous neurogenesis and synaptic remodelling throughout adult life in primary and secondary olfactory centres, its precisely ordered synaptic network and accessibility for manipulation. After homotopic transplantation of fetal olfactory bulbs in bulbectomized neonatal rodents, newly regenerated olfactory neurons form glomeruli within the graft, and the efferent mitral/tufted cells of the transplant innervate the host brain, terminating in higher olfactory centres. However, the synaptic connections of the transplanted relay neurons within the graft and/or host's olfactory centres could not be characterized mainly because of lack of suitable cell-specific markers for these neurons. In this study, we have used olfactory bulbs from transgenic fetuses, in which the majority of the mitral/tufted cells express the bacterial enzyme beta-galactosidase, for homotopic olfactory bulb transplantation following complete unilateral bulbectomy. In the transplants, the cell bodies and terminals of the donor mitral/tufted cells were identified by beta-galactosidase histochemistry and immunocytochemistry at both light and electron microscope levels. We demonstrate that transplanted relay neurons re-establish specific synaptic connections with host neurons of the periphery, source of the primary signal and central nervous system, thereby providing the basis for a functional recovery in the lesioned olfactory system.

Visualization of beta-galactosidase by enzyme and immunohistochemistry in the olfactory bulb of transgenic mice carrying the LacZ transgene.

In the olfactory bulb (OB) of a transgenic mouse line that carries the bacterial LacZ gene under the control of the 5'-regulatory region of the GAD67 gene, expression of the beta-galactosidase was confined almost exclusively to the non-GABAergic mitral and tufted cells. By light microscopy, enzyme histochemistry showed strong staining in the cell bodies and faint diffuse staining in the axons and dendrites. With immunohistochemistry for beta-galactosidase the entire cytoplasm, including the axons and dendrites, was strongly stained. By electron microscopy, beta-galactosidase enzyme histochemistry resulted in a submicroscopic reaction product that was diffusely distributed in the cytoplasm of neurons. In addition, large deposits of the reaction product were also seen attached to the cytoplasmic side of the membranes. In contrast, when the intracellular localization of beta-galactosidase was determined by immunohistochemistry, homogeneous cytoplasmic staining was obtained that filled the entire cytoplasm including the terminal dendrites and fine axons. Therefore, synaptic contacts of the beta-galactosidase-positive output neurons with other beta-galactosidase-negative neuronal cells were readily recognized in the OB. As we demonstrated, transgenic mouse lines expressing the LacZ reporter gene in a well-defined neuronal subpopulation can be used to follow beta-galactosidase-positive neurons and to directly identify their synaptic connections.

[Use of flour from sunflower oil cake in the biosynthesis of antigungal antibiotics]. / Ispol'zovanie muki iz shrota podsolnechnika pri biosinteze protivogribkovykh antibiotikov.

The possibility of replacing soybean meal and corn-steep liquor by food wastes of the oilpress industry and the meal of the sunflower oil cake in particular is discussed as applied to the fermentation media for production of antifungal antibiotics, such as levorin, mycoheptin, amphotericin. The studies showed that replacement of soybean meal by sunflower oil cake meal with simultaneous increasing of the amount of carbohydrates in the medium increased the levorin levels by 60--70 per cent as compared to the media used at present. When soybean meal and corn-steep liquor were simultaneously replaced by sunflower oil cake meal in amounts of 3--4 per cent the levels of mycoheptin in the fermentation broth increased by 30--65 per cent respectively. Replacement of soybean meal and corn-steep liquor by 3 per cent of sunflow oil cake meal in the medium used presently increased the amphotericin levels by 27 percent as compared to the control. Therefore, sunflower oil cake meal is a substitute of full value for soybean meal and corn-steep liquor in the fermentation media for production of antifungal antibiotics.