scholarly journals Assessment of Multilocus Sequence Analysis (MLSA) for Identification of Candidatus Liberibacter Solanacearum from Different Host Plants in Spain

2020 ◽  
Vol 8 (9) ◽  
pp. 1446
Author(s):  
Ana Ruiz-Padilla ◽  
Cristina Redondo ◽  
Adrián Asensio ◽  
Jerson Garita-Cambronero ◽  
Carmen Martínez ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Disease Outbreaks ◽  
Housekeeping Genes ◽  
Multilocus Sequence Analysis ◽  
Complete Analysis ◽  
Evolutionary Divergence ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Candidatus Liberibacter ◽  
Potato Crops

Liberibacter is a bacterial group causing different diseases and disorders in plants. Among liberibacters, Candidatus Liberibacter solanaceraum (CLso) produces disorders in several species mainly within Apiaceae and Solanaceae families. CLso isolates are usually grouped in defined haplotypes according to single nucleotide polymorphisms in genes associated with ribosomal elements. In order to characterize more precisely isolates of CLso identified in potato in Spain, a Multilocus Sequence Analysis (MLSA) was applied. This methodology was validated by a complete analysis of ten housekeeping genes that showed an absence of positive selection and a nearly neutral mechanism for their evolution. Most of the analysis performed with single housekeeping genes, as well as MLSA, grouped together isolates of CLso detected in potato crops in Spain within the haplotype E, undistinguishable from those infecting carrots, parsnips or celery. Moreover, the information from these housekeeping genes was used to estimate the evolutionary divergence among the different CLso by using the concatenated sequences of the genes assayed. Data obtained on the divergence among CLso haplotypes support the hypothesis of evolutionary events connected with different hosts, in different geographic areas, and possibly associated with different vectors. Our results demonstrate the absence in Spain of CLso isolates molecularly classified as haplotypes A and B, traditionally considered causal agents of zebra chip in potato, as well as the uncertain possibility of the present haplotype to produce major disease outbreaks in potato that may depend on many factors that should be further evaluated in future works.

Multilocus Sequence Analysis and Copper Ion Resistance Detection of 60 Xanthomonas arboricola pv. juglandis Isolates from China

Plant Disease ◽  
2021 ◽  
Author(s):  
Benzhong Fu ◽  
Jieqian Zhu ◽  
Conard Lee ◽  
Lihua Wang
Keyword(s):  
Sequence Analysis ◽  
Copper Ion ◽  
Housekeeping Genes ◽  
Threshold Value ◽  
Pcr Primers ◽  
Multilocus Sequence Analysis ◽  
Nucleotide Polymorphisms ◽  
Specific Pcr ◽  
Resistant Strains ◽  
Xanthomonas Arboricola

Walnut bacterial blight caused by Xanthomonas arboricola pv. juglandis (Xaj) has serious repercussions for walnut production around the world. Between 2015 and 2017, disease samples were collected from six counties (Danjiangkou, Baokang, Suizhou, Shennongjia, Zigui, and Xingshan) in Hubei province, China. Fifty-nine Xaj strains were identified by morphology and specific PCR primers from 206 isolates. The genetic diversity of 60 Xaj strains (59 from Hubei plus one from Beijing) was evaluated by Multilocus Sequence Analysis (MLST), and their resistance to copper ion (Cu2+) treatment was determined. A Neighbor Joining phylogenetic dendrogram was constructed based on four sequences of housekeeping genes (atpD-dnaK-glnA-gyrB). Two groups of strains were identified whose clustering was consistent with that of glnA. The minimal inhibitory concentration of copper ion on representative Xaj strain DW3F3 (the first genome sequenced Xaj from China) was 115 μg/ml. Setting the copper resistant threshold value to 125 μg/ml, 47 and 13 strains were considered sensitive and resistant to Cu2+, respectively. Furthermore, five strains showed Cu2+ resistance at 270 μg/ml. Compared to the copB from sensitive strains, the copB gene in resistant strains had a 15-bp insertion and eight scattered single nucleotide polymorphisms. Interestingly, the clustering based on MLSA was distinct between Xaj copper ion resistant and sensitive strains.

scholarly journals Multilocus sequence analysis of Ensifer and related taxa

2007 ◽  
Vol 57 (3) ◽  
pp. 489-503 ◽  
Author(s):  
Miet Martens ◽  
Manuel Delaere ◽  
Renata Coopman ◽  
Paul De Vos ◽  
Monique Gillis ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
16S Rrna ◽  
Housekeeping Genes ◽  
16S Rrna Genes ◽  
New Combination ◽  
Rrna Genes ◽  
Multilocus Sequence Analysis ◽  
Bootstrap Support ◽  
Rrna Gene ◽  
Separate Species

Multilocus sequence analysis (MLSA) was performed on representatives of Ensifer (including species previously assigned to the genus Sinorhizobium) and related taxa. Neighbour-joining (NJ), maximum-parsimony (MP) and maximum-likelihood (ML) phylogenies of dnaK, gltA, glnA, recA, thrC and 16S rRNA genes were compared. The data confirm that the potential for discrimination of Ensifer species is greater using MLSA of housekeeping genes than 16S rRNA genes. In incongruence-length difference tests, the 16S rRNA gene was found to be significantly incongruent with the other genes, indicating that this gene should not be used as a single indicator of relatedness in this group. Significant congruence was detected for dnaK, glnA and thrC. Analyses of concatenated sequences of dnaK, glnA and thrC genes yielded very similar NJ, MP and ML trees, with high bootstrap support. In addition, analysis of a concatenation of all six genes essentially produced the same result, levelling out potentially conflicting phylogenetic signals. This new evidence supports the proposal to unite Ensifer and Sinorhizobium in a single genus. Support for an alternative solution preserving the two genera is less strong. In view of the opinions expressed by the Judicial Commission, the name of the genus should be Ensifer, as proposed by Young [Young, J. M. (2003). Int J Syst Evol Microbiol 53, 2107–2110]. Data obtained previously and these new data indicate that Ensifer adhaerens and ‘Sinorhizobium morelense’ are not heterotypic synonyms, but represent separate species. However, transfer to the genus Ensifer is not possible at present because the species name is the subject of a pending Request for an Opinion, which would affect whether a novel species in the genus Ensifer or a new combination based on a basonym would be created.

scholarly journals Description and Validation of a Novel Real-Time RT-PCR Enterovirus Assay

2008 ◽  
Vol 54 (2) ◽  
pp. 406-413 ◽  
Author(s):  
Weston C Hymas ◽  
Wade K Aldous ◽  
Edward W Taggart ◽  
Jeffery B Stevenson ◽  
David R Hillyard
Keyword(s):  
Sequence Analysis ◽  
Single Nucleotide Polymorphisms ◽  
Real Time ◽  
Clinical Samples ◽  
Nucleotide Polymorphisms ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Single Nucleotide ◽  
The Real ◽  
Nontranslated Region

Abstract Background: Enteroviruses are a leading cause of aseptic meningitis in adult and pediatric populations. We describe the development of a real-time RT-PCR assay that amplifies a small target in the 5′ nontranslated region upstream of the classical Rotbart enterovirus amplicon. The assay includes an RNA internal control and incorporates modified nucleotide chemistry. Methods: We evaluated the performance characteristics of this design and performed blinded parallel testing on clinical samples, comparing the results with a commercially available RT-PCR assay (Pan-Enterovirus OligoDetect kit) that uses an enzyme immunoassay–like plate end detection. Results: We tested 778 samples and found 14 discrepant samples between the 2 assays. Of these, the real-time assay detected 6 samples that were negative by the OligoDetect kit, 5 of which were confirmed as positive by sequence analysis using an alternative primer set. Eight discrepant samples were positive by the OligoDetect kit and real-time negative, with 6 confirmed by sequencing. Overall, detection rates of 97% and 96% were obtained for the OligoDetect kit and real-time assays, respectively. Sequence analysis revealed the presence of a number of single nucleotide polymorphisms in the targeted region. The comparative sensitivities of the 2 assays were equivalent, with the limit of detection for the real-time assay determined to be approximately 430 copies per milliliter in cerebrospinal fluid. Conclusions: This novel real-time enterovirus assay is a sensitive and suitable assay for routine clinical testing. The presence of single nucleotide polymorphisms can affect real-time PCR assays.

scholarly journals A New Borrelia Species Defined by Multilocus Sequence Analysis of Housekeeping Genes

2009 ◽  
Vol 75 (16) ◽  
pp. 5410-5416 ◽  
Author(s):  
Gabriele Margos ◽  
Stephanie A. Vollmer ◽  
Muriel Cornet ◽  
Martine Garnier ◽  
Volker Fingerle ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Lyme Borreliosis ◽  
Phylogenetic Trees ◽  
Evolutionary History ◽  
Housekeeping Genes ◽  
Multilocus Sequence Analysis ◽  
Species Status ◽  
Borrelia Garinii ◽  
Analysis Scheme ◽  
History Of

ABSTRACT Analysis of Lyme borreliosis (LB) spirochetes, using a novel multilocus sequence analysis scheme, revealed that OspA serotype 4 strains (a rodent-associated ecotype) of Borrelia garinii were sufficiently genetically distinct from bird-associated B. garinii strains to deserve species status. We suggest that OspA serotype 4 strains be raised to species status and named Borrelia bavariensis sp. nov. The rooted phylogenetic trees provide novel insights into the evolutionary history of LB spirochetes.

scholarly journals Multilocus Sequence Analysis Reveals Genetic Diversity in Xanthomonads Associated With Poinsettia Production

Plant Disease ◽  
2015 ◽  
Vol 99 (6) ◽  
pp. 874-882 ◽  
Author(s):  
W. Rockey ◽  
N. Potnis ◽  
S. Timilsina ◽  
J. C. Hong ◽  
G. E. Vallad ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Leaf Spot ◽  
Reference Strain ◽  
Housekeeping Genes ◽  
Causal Agent ◽  
Multilocus Sequence Analysis ◽  
Bacterial Leaf Spot ◽  
The Family ◽  
Systemic Infections ◽  
Spot Formation

Xanthomonas axonopodis pv. poinsettiicola is traditionally identified as the primary causal agent of bacterial leaf spot on poinsettia (family Euphorbiaceae). Sixty-seven strains of xanthomonads isolated from lesions associated with several species within the family Euphorbiaceae were collected over a 64-year period. The pathogenicity of these strains was compared on several potential hosts and they were analyzed by multilocus sequence analysis (MLSA) using six housekeeping genes. The 67 Xanthomonas strains associated with poinsettia production were separated into three distinct clades based on MLSA. The first clade identified contained the X. axonopodis pv. poinsettiicola reference strain (LMG849PT). A second clade was more closely related to X. hortorum pv. pelargonii (LMG7314PT) and the third clade contained the X. codiaei type strain (LMG8678T). This analysis indicated that there may also be other closely related pathovars or species of Xanthomonas that can infect poinsettia. Strains from the three clades could not be distinguished by symptoms or virulence on poinsettia plants. Strains capable of infecting geranium were found in all three clades, although the extent of leaf spot formation and number of systemic infections were significantly less than those produced by X. hortorum pv. pelargonii strains, typically the main causal agent of bacterial leaf spot on geranium. Clade III also contained strains isolated from zebra plant (Aphelandra squarrosa, family Acanthaceae), which is a newly recognized host for X. codiaei and X. axonopodis pv. poinsettiicola. Xanthomonas leaf spot is a serious threat to poinsettia production that can be caused by several Xanthomonas spp. that can infect different ornamental plant hosts. It is imperative that growers maintain a strict sanitation program because reservoirs of inoculum can occur on a number of ornamental hosts.

scholarly journals Multilocus Sequence Analysis of Xanthomonads Causing Bacterial Spot of Tomato and Pepper Plants Reveals Strains Generated by Recombination among Species and Recent Global Spread of Xanthomonas gardneri

2014 ◽  
Vol 81 (4) ◽  
pp. 1520-1529 ◽  
Author(s):  
Sujan Timilsina ◽  
Mustafa O. Jibrin ◽  
Neha Potnis ◽  
Gerald V. Minsavage ◽  
Misrak Kebede ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Sequence Analysis ◽  
Housekeeping Genes ◽  
Global Distribution ◽  
Multilocus Sequence Analysis ◽  
Regional Differentiation ◽  
Bacterial Spot ◽  
Content Type ◽  
Global Spread ◽  
Pathogen Populations

ABSTRACTFourXanthomonasspecies are known to cause bacterial spot of tomato and pepper, but the global distribution and genetic diversity of these species are not well understood. A collection of bacterial spot-causing strains from the Americas, Africa, Southeast Asia, and New Zealand were characterized for genetic diversity and phylogenetic relationships using multilocus sequence analysis of six housekeeping genes. By examining strains from different continents, we found unexpected phylogeographic patterns, including the global distribution of a single multilocus haplotype ofX. gardneri, possible regional differentiation inX. vesicatoria, and high species diversity on tomato in Africa. In addition, we found evidence of multiple recombination events betweenX. euvesicatoriaandX. perforans.Our results indicate that there have been shifts in the species composition of bacterial spot pathogen populations due to the global spread of dominant genotypes and that recombination between species has generated genetic diversity in these populations.

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