Effects of intra-accumbal or intra-prefrontal cortex microinjections of adenosine 2A receptor ligands on responses to cocaine reward and seeking in rats.

RATIONALE AND OBJECTIVES: Many studies indicated that adenosine via its A2A receptors influences the behavioral effects of cocaine by modulating dopamine neurotransmission. The hypothesis was tested that A2A receptors in the nucleus accumbens (NAc) or the prefrontral cortex (PFc) may modulate cocaine reward and/or cocaine seeking behavior in rats. METHODS: The effects of local bilateral microinjections of the selective A2A receptor agonist CGS 21680 or the A2A receptor antagonists KW 6002 and SCH 58261 were investigated on cocaine self-administration on reinstatement of cocaine seeking. RESULTS: The intra-NAc shell, but not intra-infralimbic PFc, administration of CGS 21680 significantly reduced the number of active lever presses and the number of cocaine (0.25 mg/kg) infusions. However, tonic activation of A2A receptors located in the NAc or PFc did not play a role in modulating the rewarding actions of cocaine since neither KW 6002 nor SCH 58261 microinjections altered the cocaine (0.5 mg/kg) infusions. The intra-NAc but not intra-PFc microinjections of CGS 21680 dose- dependently attenuated the reinstatement of active lever presses induced by cocaine (10 mg/kg, i.p.) and the drug-associated combined conditioned stimuli using the subthreshold dose of cocaine (2.5 mg/kg, i.p.). On the other hand, the intra-NAc pretreatment with SCH 58261, but not with KW 6002, given alone evoked reinstatement of cocaine seeking behavior. CONCLUSION: The results strongly support the involvement of accumbal shell A2A receptors as a target, the activation of which exerts an inhibitory control over cocaine reward and cocaine seeking.

Taxonomic relatedness between Pectobacterium carotovorum subsp. carotovorum, Pectobacterium carotovorum subsp. odoriferum and Pectobacterium carotovorum subsp. brasiliense subsp. nov.

AIMS: Pectobacterium carotovorum is a heterogeneous species consisting of two named subspecies, P. carotovorum subsp. carotovorum and P. carotovorum subsp. odoriferum. A third subspecies, P. carotovorum subsp. brasiliense, was previously proposed. The study aimed to confirm the subspecies status and validate the proposed name of P. carotovorum subsp. brasiliense using a novel and standard microbial taxonomy. METHODS AND RESULTS: DNA-DNA hybridization confirmed that P. carotovorum subsp. brasiliense is a different species from P. wasabiae, P. betavasculorum and P. atrosepticum, with 28, 35 and 55% similarity values, respectively, but is a member of the P. carotovorum species with 73-77% similarity values. Sequencing the entire 16S rRNA gene of two polymorphic copies from strains of each of the P. carotovorum subspecies demonstrated that the average 16S rRNA gene sequence diversity between P. carotovorum subsp. brasiliense and P. carotovorum subsp. carotovorum was lower than the maximum genetic distances between two sequence types obtained from the same strain. Multilocus sequence analysis based on eight housekeeping genes (mtlD, acnA, icdA, mdh, pgi, gabA, proA and rpoS) differentiated the subspecies and delineated two P. carotovorum subsp. brasiliense clades. CONCLUSION: Pectobacterium carotovorum subsp. brasiliense clade I was comprised of strains isolated from Brazil and Peru, while clade II included strains from Asia, North America and Europe. Strains in clade I but not clade II were phenotypically consistent with the original description of P. carotovorum subsp. brasiliense in that they produced reducing substances from sucrose and acid from α-methyl glucoside. The type strain for P. carotovorum subsp. brasiliense 212(T) (= LMG2137(T) = IBSBF1692(T) = CFBP6617(T) ) was previously designated. The GC mol content of the type strain is 51·7%. SIGNIFICANT AND IMPACT OF THE STUDY: the study introduces a full description for the strains belonging to the two different clades assigned to P. carotovorum subsp. brasiliense.

The importance of the adenosine A(2A) receptor-dopamine D(2) receptor interaction in drug addiction.

Drug addiction is a serious brain disorder with somatic, psychological, psychiatric, socio-economic and legal implications in the developed world. Illegal (e.g., psychostimulants, opioids, cannabinoids) and legal (alcohol, nicotine) drugs of abuse create a complex behavioral pattern composed of drug intake, withdrawal, seeking and relapse. One of the hallmarks of drugs that are abused by humans is that they have different mechanisms of action to increase dopamine (DA) neurotransmission within the mesolimbic circuitry of the brain and indirectly activate DA receptors. Among the DA receptors, D(2) receptors are linked to drug abuse and addiction because their function has been proven to be correlated with drug reinforcement and relapses. The recognition that D(2) receptors exist not only as homomers but also can form heteromers, such as with the adenosine (A)(2A) receptor, that are pharmacologically and functionally distinct from their constituent receptors, has significantly expanded the range of potential drug targets and provided new avenues for drug design in the search for novel drug addiction therapies. The aim of this review is to bring current focus on A(2A) receptors, their physiology and pharmacology in the central nervous system, and to discuss the therapeutic relevance of these receptors to drug addiction. We concentrate on the contribution of A(2A) receptors to the effects of different classes of drugs of abuse examined in preclinical behavioral experiments carried out with pharmacological and genetic tools. The consequences of chronic drug treatment on A(2A) receptor-assigned functions in preclinical studies are also presented. Finally, the neurochemical mechanism of the interaction between A(2A) receptors and drugs of abuse in the context of the heteromeric A(2A)-D(2) receptor complex is discussed. Taken together, a significant amount of experimental analyses provide evidence that targeting A(2A) receptors may offer innovative translational strategies for combating drug addiction.

First Report of Bacterial Wilt of Tomato (Solanum lycopersicum) Caused by Ralstonia solanacearum in Benin.

In June 2004, wilted tomatoes with no foliar yellowing were observed in Ouègbo, Atlantique District, Benin. The cut tomato stems released whitish bacterial ooze. Longitudinal sections of most stems showed brown vascular discoloration. Twenty symptomatic tomato plants were collected from 10 fields and exported to the Institute of Plant Disease and Plant Protection, Leibniz Universität Hannover, Germany. Bacteria were isolated on triphenyl tetrazolium chloride (TTC) medium (2) and three of the nine bacterial isolates that resembled Ralstonia solanacearum (colonies with red center and whitish periphery) and reference strain ToUdk (race 1 biovar 3; N. Thaveechai, Kasetsart University, Bangkok, Thailand) were used for pathogenicity tests. Five 4-week-old tomato plants cv. Tohounvi, grown in individual plastic pots (14 × 16 cm) containing sterilized field soil, were inoculated with each of the four isolates individually by soil drenching with 30 ml of the test cultures at 108 CFU/ml. Control plants were treated with 30 ml of sterile water. All plants were incubated in a glasshouse at 30°C. All plants inoculated with the isolates from Benin wilted 4 days after inoculation with symptoms similar to those observed in the field. Plants inoculated with the reference strain wilted 7 to 11 days after inoculation. Control plants treated with water remained healthy. R. solanacearum was recovered from the 20 symptomatic plants on TTC medium. The identity of the strains in comparison with the reference strain was confirmed by PCR with species-specific primers 759/760, which produced a single 281-bp fragment (3). Because similar symptoms were being increasingly reported by farmers across Benin and linked with reduced tomato yields, a disease survey was undertaken by IITA in 2006 and 2007. Wilted tomato plants were found across all agro-ecological zones of Benin (3 to 72% of plants per field). Isolates were recovered from the southeastern districts of Adja-Ouèrè, Sakété, Adjohoun, and Dangbo, the southwestern districts of Klouékanmè and Athiémé, the southern districts of Toffo and Bohicon, the central districts of Dassa and Savè, and the northern districts of Malanville and Karimama. Identification of R. solanacearum was confirmed following inoculation of tomato, production of characteristic wilting symptoms, recovery of the pathogen on TTC medium, and positive identification with ELISA kits (Pathoscreen Rs; Agdia Inc., Elkhart, IN). To our knowledge, this is the first report of R. solanacearum infecting tomato in Benin. Tomato is the most cultivated vegetable crop in Benin and important to the livelihood of many people in peri-urban and rural areas. Understanding that the cause of the observed crop losses is R. solanacearum may lead to implementation of management strategies such as deployment of disease-resistant cultivars or grafting tomatoes onto bacterial wilt-resistant rootstocks (1). References: (1) P. Aggarwal et al. Indian J. Agric. Sci. 78:379, 2008. (2) A. Kelman. Phytopathology 44:693, 1954. (3) N. Opina et al. Asian Pac. J. Mol. Biol. Biotechnol. 5:19, 1997.

First Report of Bacterial Speck of Tomato Caused by Pseudomonas syringae pv. tomato in Tanzania.

In April 2004, there was a serious outbreak of a tomato (Lypersicon esculentum Mill.) leaf spot disease in Mgeta, Mvomero District of Tanzania. The disease was characterized by lesions on green tomato fruits that were small, sunken, and black and were surrounded by darker green haloes. Lesions on ripe tomato fruits were dark brown to black, superficial, and measured approximately 1 to 2 mm in diameter. On the leaves, lesions were small, black, and surrounded by chlorotic (yellow) haloes. In some cases, the specks coalesced to form large lesions on older leaves. Black lesions were also observed on stems and petioles. A disease survey of selected tomato-producing areas in Arusha, Dodoma, Iringa, and Morogoro regions of Tanzania during 2004 and 2005 revealed that the disease was widespread in farmers' fields in all areas surveyed. Disease incidence was approximately 80%, while severity, rated on the scale of Chambers and Merriman (1), ranged from moderate (11 to 40 lesions per plant) to severe (>40 lesions per plant). A bacterium that produced a greenish, diffusible pigment on King's medium B was consistently isolated from lesions on tomato fruits collected from the fields in all the surveyed areas. All 56 isolates obtained were gram negative, oxidase negative, and fluoresced on King's medium B under UV light. None utilized phenylethylamine as the sole carbon source, while three isolates utilized i-erythritol and lactulose. Biolog analysis of the isolates, along with two reference strains of P. syringae pv. tomato (Pst CEP-3 from Sokoine University of Agriculture, Tanzania and Pst BB6 [Race 1] from Göttinger Sammlung Phytopathogener Bakterien, Göttingen, Germany) identified them as P. syringae pv. tomato, with similarity indices of 0.518 to 0.933. They also were positively identified as P. syringae pv. tomato by repetitive sequence-based-PCR (2,3) and fragment length polymorphism analysis. Pathogenicity of the strains was confirmed by spraying 35-day-old tomato seedlings (cv. Tanya) with suspensions of the isolates at a concentration of 108 CFU ml-1 of sterile water. After approximately 72 h, small, water-soaked, dark brown lesions similar to those observed on the field plants were observed on leaves of all the inoculated tomato seedlings. There were no symptoms on control plants. The bacterium was reisolated from the infected plants and identified as P. syringae pv. tomato, in accordance with Koch's postulates. To our knowledge, this is the first report of the occurrence of tomato bacterial speck in Tanzania. References: (1). S. C. Chambers and P. R. Merriman. Aust. J. Agric. Res. 26:657, 1975. (2). F. J. Louws et al. Appl. Environ. Microbiol. 60:2286, 1994. (3). M. Zaccardelli et al. Eur. J. Plant Pathol. 111:85, 2005.

Identification of Pathotypes of Xanthomonas axonopodis pv. manihotis in Africa and Detection of Quantitative Trait Loci and Markers for Resistance to Bacterial Blight of Cassava.

ABSTRACT Cassava suffers from bacterial blight attack in all growing regions. Control by resistance is unstable due to high genotype-environment interactions. Identifying genes for resistance to African strains of Xanthomonas axonopodis pv. manihotis can support breeding efforts. Five F(1) cassava genotypes deriving from the male parent 'CM2177-2' and the female parent 'TMS30572' were used to produce 111 individuals by backcrossing to the female parent. In all, 16 genotypes among the mapping population were resistant to stem inoculation by four strains of X. axonopodis pv. manihotis from different locations in Africa, and 19 groups with differential reactions to the four strains were identified, suggesting that the strains represent different pathotypes. Four genotypes were resistant to leaf inoculation, and three were resistant to both stem and leaf inoculations. Genotypes with susceptible, moderately resistant, and resistant reactions after leaf and stem inoculation partly differed in their reactions on leaves and stems. Based on the genetic map of cassava, single-markeranalysis of disease severity after stem-puncture inoculation was performed. Eleven markers were identified, explaining between 16 and 33.3% of phenotypic variance of area under disease progress curve. Five markers on three and one linkage groups from the female- and male-derived framework of family CM8820, respectively, seem to be weakly associated with resistance to four strains of X. axonopodis pv. manihotis. Based on the segregation of alleles from the female of family CM8873, one marker was significantly associated with resistance to two X. axonopodis pv. manihotis strains, GSPB2506 and GSPB2511, whereas five markers were not linked to any linkage group. The quantitative trait loci (QTL) mapping results also suggest that the four African strains belong to four different pathotypes. The identified pathotypes should be useful for screening for resistance, and the QTL and markers will support breeding for resistance.

Development of a New Semiselective Medium for Isolating Xanthomonas campestris pv. manihotis from Plant Material and Soil.

ABSTRACT An effective control for bacterial blight of cassava (Manihot esculenta), caused by Xanthomonas campestris pv. manihotis, requires the use of non-contaminated cuttings and seeds. Using classical agar plating techniques for screening planting material for contamination has not been very successful because of the lack of a reliable semiselective agar medium. The pathogen grows slowly on general plating media and is easily overgrown by saprophytic bacteria during isolation from diseased plants. In an effort to develop a semiselective medium, the utilization of several carbon and nitrogen sources was studied. Results of these tests provided information used to design a basal medium allowing good growth of the target organism while suppressing growth of several common saprophytes. Additional selectivity was achieved by incorporating three antibiotics into the basal medium. The new semiselective agar medium, designated cefazolin trehalose agar (CTA) medium, contained (per liter) 3.0 g of K(2)HPO(4), 1.0 g of NaH(2)PO(4), 0.3 g of MgSO(4).7H(2)O, 1.0 g of NH(4)Cl, 9.0 g of D(+)-trehalose, 1.0 D(+)-glucose, 1.0 g of yeast extract, 0.025 g of cefazolin, 0.0012 g of lincomycin, 0.0025 g of phosphomycin, 0.25 g of cycloheximide, and 14.0 g of agar. In comparison to a starch-based semiselective medium (SXM), plating efficiencies using pure cultures of 10 strains of X. campestris pv. manihotis were significantly higher on CTA, with an average of 85 and 50%, respectively. Likewise, isolation and recovery of X. campestris pv. manihotis from infected cassava leaves and contaminated soil were much higher on CTA than on SXM agar. When X. campestris pv. manihotis occurs in high concentrations in diseased tissue, the standard yeast trehalose glucose agar medium supplemented with 250 mug of cycloheximide per ml appears to be satisfactory. The newly developed CTA medium should prove useful for control strategies to identify and remove infected planting material of cassava, as well as for basic ecological studies of the pathogen.