Cassava diseases caused by Xanthomonas phaseoli pv. manihotis and Xanthomonas cassavae

Xanthomonas phaseoli pv. manihotis (Xpm) and X. cassavae (Xc) are two bacterial pathogens attacking cassava. Cassava bacterial blight (CBB) is a systemic disease caused by Xpm, which might have dramatic effects on plant growth and crop production. Cassava bacterial necrosis is a nonvascular disease caused by Xc with foliar symptoms similar to CBB, but its impacts on the plant vigour and the crop are limited. In this review, we describe the epidemiology and ecology of the two pathogens, the impacts and management of the diseases, and the main research achievements for each pathosystem. Because Xc data are sparse, our main focus is on Xpm and CBB.     

Integron variability in Xanthomonas arboricola pv. juglandis and Xanthomonas arboricola pv. pruni strains

The integron platform and the gene cassette arrays of 34 Xanthomonas arboricola pv. juglandis and of 47 Xanthomonas arboricola pv. pruni strains isolated from different geographical areas were screened to check their variability. Genetic variability of the strains was also tested by means of BOX-PCR. For two representative strains of the two pathovars, the integrase gene intI and part of the flanking gene ilvD were also cloned and sequenced. Whereas X. a. pv. pruni strains did not show relevant variability, six X. a. pv. juglandis strains isolated in Australia showed some differences in the gene sequences. The CLUSTALW algorithm indicated that the majority of the X. a. pv. juglandis strains are closely related to X. a. pv. pruni, whereas the X. a. pv. juglandis strains isolated in Australia were more similar to Xanthomonas hortorum pv. pelargonii. Similarly, the gene cassette array pattern of the Australian strains, as well as that of the oldest strain maintained in culture, was different from the other strains. Also, three X. a. pv. pruni strains showed a different cassette array pattern when compared with the majority of other strains but no relationships with geographical area of isolation or host plant was revealed. This study confirmed that in addition to species, integrons may generate diversity also within two X. arboricola pathovars.     

Phenotypic diversity of Xanthomonas sp. mangiferaeindicae

Carbohydrate utilization profiles by means of the API (Appareils et Procédés d'Identification) system and sensitivity to antibiotics and heavy metal salts of 68 Xanthomonas sp. mangiferaeindicae strains isolated in nine countries from mango (Mangifera indica L.) and other genera of the Anacardiaceae were examined to assess the variability of the taxon. The strains could be separated into 10 groups according to Ward clustering. Apigmented strains isolated from the pepper tree [syn. Brazilian pepper] (Schinus terebenthifolius Raddi) could not be clearly differentiated from most apigmented strains isolated from mango. Yellow-pigmented strains isolated from mango in Brazil and Reunion Island, apigmented strains isolated from mango in Brazil and from ambarella in the French West Indies, clustered in distinct groups. The results are consistent with those of other studies, based on isozyme analysis of esterase, phosphoglucomutase and superoxide dismutase, and hrp-RFLP analysis; they indicate the need for a comprehensive taxonomic evaluation of xanthomonads associated with Anacardiaceae.     

Molecular characterization of copper resistance genes from Xanthomonas citri subsp. citri and Xanthomonas alfalfae subsp. citrumelonis

Copper sprays have been widely used for control of endemic citrus canker caused by Xanthomonas citri subsp. citri in citrus-growing areas for more than 2 decades. Xanthomonas alfalfae subsp. citrumelonis populations were also exposed to frequent sprays of copper for several years as a protective measure against citrus bacterial spot (CBS) in Florida citrus nurseries. Long-term use of these bactericides has led to the development of copper-resistant (Cu(r)) strains in both X. citri subsp. citri and X. alfalfae subsp. citrumelonis, resulting in a reduction of disease control. The objectives of this study were to characterize for the first time the genetics of copper resistance in X. citri subsp. citri and X. alfalfae subsp. citrumelonis and to compare these organisms to other Cu(r) bacteria. Copper resistance determinants from X. citri subsp. citri strain A44(pXccCu2) from Argentina and X. alfalfae subsp. citrumelonis strain 1381(pXacCu2) from Florida were cloned and sequenced. Open reading frames (ORFs) related to the genes copL, copA, copB, copM, copG, copC, copD, and copF were identified in X. citri subsp. citri A44. The same ORFs, except copC and copD, were also present in X. alfalfae subsp. citrumelonis 1381. Transposon mutagenesis of the cloned copper resistance determinants in pXccCu2 revealed that copper resistance in X. citri subsp. citri strain A44 is mostly due to copL, copA, and copB, which are the genes in the cloned cluster with the highest nucleotide homology (≥ 92%) among different Cu(r) bacteria.     

Phenotypic diversity of Xanthomonas sp. mangiferaeindicae

Carbohydrate utilization profiles by means of the API (Appareils et Procédés d'Identification) system and sensitivity to antibiotics and heavy metal salts of 68 Xanthomonas sp. mangiferaeindicae strains isolated in nine countries from mango ( Mangifera indica L.) and other genera of the Anacardiaceae were examined to assess the variability of the taxon. The strains could be separated into 10 groups according to Ward clustering. Apigmented strains isolated from the pepper tree [syn. Brazilian pepper] ( Schinus terebenthifolius Raddi) could not be clearly differentiated from most apigmented strains isolated from mango. Yellow-pigmented strains isolated from mango in Brazil and Reunion Island, apigmented strains isolated from mango in Brazil and from ambarella in the French West Indies, clustered in distinct groups. The results are consistent with those of other studies, based on isozyme analysis of esterase, phosphoglucomutase and superoxide dismutase, and hrp -RFLP analysis; they indicate the need for a comprehensive taxonomic evaluation of xanthomonads associated with Anacardiaceae.     

水稻白叶枯病菌和水稻细菌性条斑病菌的实时荧光PCR快速检测鉴定

成功建立了水稻白叶枯菌与水稻细菌性条斑病菌快速检测鉴定的实时荧光PCR方法。根据含铁细胞接受子基因设计两菌的通用引物PSRGF/PSRGR（扩增一个152bpDNA片段）和特异性探针（Baiprobe和Tiaoprobe）,并对13种细菌和1种植原体进行实时荧光PCR。结果表明,两个特异性探针能分别特异性检测到目标病原菌产生荧光信号而其它参考菌不产生荧光信号。检测的绝对灵敏度是30.6fg/μL质粒DNA和103CFU/mL的菌悬浮液,相当于1个细菌细胞的基因,比常规PCR电泳检测高约100倍,相对灵敏度为105CFU/mL。整个检测过程只需2h,完全闭管,降低了污染的机会,无需PCR后处理。 用这两个特异性探针分别对自然感染白叶枯菌和条斑菌的叶片DNA提取液和种子浸泡液进行实时荧光PCR,结果均可特异性检测到目标菌的存在并完全可将两种病原细菌区分开来,且只需03g叶片和10g种子。     

Immunomagnetic isolation of Xanthomonas campestris pv. pelargonii

Immunomagnetic fishing was developed as an improved procedure for increasing the bacterial target to non-target recovery ratio in suspensions containing mixtures of target and non-target organisms. A cell suspension containing the target Xanthomonas campestris pv. pelargonii and non-target organisms, is treated with rabbit polyclonal antiserum against X.c. pv. pelargonii and incubated for 1 h. The suspension is then mixed with paramagnetic iron oxide particles coated with goat anti-rabbit antibodies (immunomagnetic particles). After incubation, the polished surface of a 14 mm diameter neodymium supermagnet is placed at the air-water interace and the magnetic particles are attracted to the magnet. After all visible magnetic particles have attached to the bottom of the magnet, the magnet is dipped in sterile buffer to remove non-target organisms. The magnet with attached magnetic particles is rubbed evenly over an agar surface to dislodge the particles and attached bacteria. Conventional immunomagnetic isolation (immunomagnetic attraction) and immunomagnetic fishing were compared, for the recovery of the target organism in geranium leaf washings spiked with X.c. pv. pelargonii. With immunomagnetic attraction and immunomagnetic fishing, bacterial non-target organisms were reduced to 11.4 and 1.5% of the initial population, respectively, whereas the target was only reduced to 63.7 and 53.8%.     

Conformation of the extracellular polysaccharide of Xanthomonas campestris.

The solution conformation of the extracellular polysaccharide of the bacterium Xanthomonas campestris is examined by optical rotation, viscometry, and potentiometric titration. Measurements of optical rotation vs. temperature for solutions of the polysaccharide at low ionic strength reveal a sharp transition to a denatured structure which is reversible if sufficient salt is present. The temperature Tm at the transition midpoint increases as log (Na+) or log (Ca2+). Viscosity-temperature profiles substantiate a structural change of the polysaccharide at Tm. The intrinsic viscosity of the native molecule at zero shear rate exceeds 5000 ml/g. This high figure is indicative of a stiff chain. The viscosity of the native molecule is relatively insensitive to salt, whereas the denatured molecule collapses if salt is present. Hydrogen-ion titration shows that the pKapp of the COO- groups of the polymer decreases from 3.2 in 0.01 M NaC1 to 2.6 in 0.2 M NaC1. All these data suggest that the native polysaccharide possesses ordered secondary structure stabilized by nonionic interactions outweighing the repulsion between adjacent COO- groups.     

Efficient transformation of phytopathogenic strains of Xanthomonas species.

Efficient transformation and conjugation systems for use in phytopathogenic strains of Xanthomonas species were developed with a dual-function plasmid vector, pBXC12, which was constructed from a newly isolated Xanthomonas citri plasmid, pXCL6, and pBR328. By using this system, pBR328 could also transform the same strains of Xanthomonas. The systems make useful cloning vectors for the study of genes involved in the plant pathogenesis of this species.     

Culture medium for Xanthomonas campestris pv. oryzae

Y uan , W. 1990. Culture medium for Xanthomonas campestris pv. oryzae. Journal of Applied Bacteriology 69 , 798–805.
Studies on nutrient requirements of four Chinese strains of Xanthomonas campestris pv. oryzae in a modified Watanabe's medium led to the development of a new synthetic medium containing sucrose, sodium glutamate, methionine, KH₂PO₄, NH₄C1 and iron chelated with EDTA. The concentration of each ingredient was optimized based on the number of colonies and time required for their appearance. Various concentrations of some nutrients were compared based upon their effects on growth of the pathogen strains and 34 contaminants from rice materials. Tryp-tone enhanced the growth of X. c. oryzae more than that of many contaminants, including Erwinia herbicola . Peptone stimulated growth of X. c. oryzae without promoting excessive contamination. When compared with other media used for X. c. oryzae , the new culture medium enriched with tryptone and peptone gave the highest recovery and earliest appearance of colonies of Chinese strains of this bacterium.     

A third restriction endonuclease from Xanthomonas malvacearum.

An additional sequence-specific endonuclease, XmaIII, has been partially purified from Xanthomonas malvacearum. XmaIII recognizes ten cleavage sites in adenovirus 2 DNA, two sites in bacteriophage lambda and no site in either simian virus 40 DNA or φX174 DNA. It recognizes the sequence

and cleaves at the sites indicated by the arrows. No other endonuclease with this particular nucleotide sequence specificity has been reported.     

Identification of Xanthomonas fragariae, Xanthomonas axonopodis pv. phaseoli, and Xanthomonas fuscans subsp. fuscans with novel markers and using a dot blot platform coupled with automatic data analysis

Phytosanitary regulations and the provision of plant health certificates still rely mainly on long and laborious culture-based methods of diagnosis, which are frequently inconclusive. DNA-based methods of detection can circumvent many of the limitations of currently used screening methods, allowing a fast and accurate monitoring of samples. The genus Xanthomonas includes 13 phytopathogenic quarantine organisms for which improved methods of diagnosis are needed. In this work, we propose 21 new Xanthomonas-specific molecular markers, within loci coding for Xanthomonas-specific protein domains, useful for DNA-based methods of identification of xanthomonads. The specificity of these markers was assessed by a dot blot hybridization array using 23 non-Xanthomonas species, mostly soil dwelling and/or phytopathogens for the same host plants. In addition, the validation of these markers on 15 Xanthomonas spp. suggested species-specific hybridization patterns, which allowed discrimination among the different Xanthomonas species. Having in mind that DNA-based methods of diagnosis are particularly hampered for unsequenced species, namely, Xanthomonas fragariae, Xanthomonas axonopodis pv. phaseoli, and Xanthomonas fuscans subsp. fuscans, for which comparative genomics tools to search for DNA signatures are not yet applicable, emphasis was given to the selection of informative markers able to identify X. fragariae, X. axonopodis pv. phaseoli, and X. fuscans subsp. fuscans strains. In order to avoid inconsistencies due to operator-dependent interpretation of dot blot data, an image-processing algorithm was developed to analyze automatically the dot blot patterns. Ultimately, the proposed markers and the dot blot platform, coupled with automatic data analyses, have the potential to foster a thorough monitoring of phytopathogenic xanthomonads.     

Phosphorylation of proteins in Xanthomonas campestris pv. oryzae

Protein phosphorylation was studied in Xanthomonas campestris pv. oryzae in vivo and in vitro. In vitro labelling showed that the protein kinases in this bacterium used both ATP and GTP as nucleotide substrates at nearly the same efficiency. At least 6 proteins were phosphorylated in vitro, including abundant species of p81, p44, and p32 with M _r of 81000, 44000, and 32000, respectively. Three types of phosphate-protein linkage were found in this bacterium: O-phosphate, N-phosphate and probably acyl phosphate. The p81 and p32 were phosphorylated at histidine. The p44 had mainly phosphoserine and a small part of phosphohistidine. The phosphorylation profile was variable depending on the growth conditions. Furthermore, by a virulent phage Xp10 infection the quantity of phosphorylation increased: for phosphohistinine more than 10-fold, and for phosphoserine about 3-fold. Thus, in this bacterium phosphorylation may be linked with a physiological regulation system and with Xp10 phage development.     

Une phytotoxine de Xanthomonas campestris pv. vasculorum

A phytotoxin from Xanthomonas campestris pv. vasculorum I. Purification and partial characterization Xanthomonas campestris pv. vasculorum, the causal agent of sugarcane disease produces, when maintained in pure culture, a host selective phytotoxin. The molecule obtained at a high purity level is a strongly acid polyalcohol.     

Status of Streptomycin Resistance Development in Xanthomonas oryzae pv. oryzae and Xanthomonas oryzae pv. oryzicola in China and their Resistance Characters

Rice leaves with bacterial blight or bacterial leaf streak symptoms were collected in southern China in 2007 and 2008. Five hundred and thirty‐four single‐colony isolates of Xanthomonas oryzae pv. oryzae and 827 single‐colony isolates of Xanthomonas oryzae pv. oryzicola were obtained and tested on plates for sensitivity to streptomycin. Four strains (0.75%) of X. oryzae pv. oryzae isolated from the same county of Province Yunnan were resistant to streptomycin, and the resistance factor (the ratio of the mean median effective concentration inhibiting growth of resistant isolates to that of sensitive isolates) was approximately 226. The resistant isolate also showed streptomycin resistance in vivo. In addition to resistant isolates, isolates of less sensitivity were also present in the population of X. oryzae pv. oryzae from Province Yunnan. However, no isolates with decreased streptomycin‐sensitivity were obtained from the population of X. oryzae pv. oryzicola. Mutations in the rpsL (encoding S12 protein) and rrs genes (encoding 16S rRNA) and the presence of the strA gene accounting for streptomycin resistance in other phytopathogens or animal and human pathogenic bacteria were examined on sensitive and resistant strains of X. oryzae pv. oryzae by polymerase chain reaction amplification and sequencing. Neither the presence of the strA gene nor mutations in the rpsL or rrs were found, suggesting that different resistance mechanisms are involved in the resistant isolates of X. oryzae pv. oryzae.     

Purification and characterization of ascamycin-hydrolysing aminopeptidase from Xanthomonas citri.

Intact obese rats were hyperinsulinaemic, had higher rates of whole-body fatty acid synthesis, higher activities of hepatic acetyl-CoA carboxylase and tyrosine aminotransferase and a higher hepatic glycogen concentration than intact lean animals. Adrenalectomy abolished all these factors of the obese phenotype. Treatment of adrenalectomized rats with corticosterone for 24 h increased the rate of whole-body fatty acid synthesis to the same extent in both phenotypes, but caused a larger increase in glycogen concentration, tyrosine aminotransferase activity and plasma insulin concentration in obese rats.