Publications

3805

IntroductionHuanglonbing (HLB) is the most serious disease of citrus in the world, associated with three non-cultivable phloem-restricted bacteria Candidatus Liberibacter asiaticus (CLas), Ca L. africanus (CLaf) and Ca L. americanus (CLam). CLas is transmitted by the Asian citrus psyllid Diaphorina citri, and has spread to several countries. The African psyllid Trioza erytreae, the vector of CLaf occurs in Africa and neighbouring islands. Only two major citrus-growing regions - Australia/New Zealand and the Mediterranean Basin - are still HLB-free in the world. However, T. erytreae has recently been introduced into continental Europe (Portugal and Spain) and has become a potential threat to citrus production. The transmission of CLas by T. erytreae had been postulated but never tested. To evaluate the risk of T. erytreae transmitting CLas, comparative transmissions of CLas by T. erytreae and D. citri were assessed.MethodsTransmission tests were performed on excised leaves and seedlings of Citrus volkameriana with different inoculation access periods (in series) for both insect species. Quantifications of bacterial titers were made in excised leaves, seedlings three and six months after inoculation and on individual insects.ResultsOur results showed that T. erytreae was able to efficiently acquire CLas. Furthermore, T. erytreae carried significantly higher bacterial titers than D. citri, and was able to efficiently transmit the bacteria to seedlings at a similar rate that D. citri highlighting the high risk of spread of the most aggressive variant of HLB (CLas) by T. erytreae in Europe.DiscussionThus, extreme precautions to prevent any entry of CLas into Europe should be adopted.

AbstractMolecular computing was used to investigate the possible causal agents of chilli crop samples showing mixed symptoms of yellow leaf curl and little leaf type diseases in the Uttar Pradesh province, India. Total genomic DNA was extracted from twenty-five samples and amplified by PCR using a universal primer pair for begomovirus and phytoplasma. Mixed infection samples show positive amplified products for begomovirus (DNA-A and betasatellite) and phytoplasma (16S rRNA and Sec A). The identified begomovirus from chilli samples was identified as a strain isolate of the previously described Chilli Leaf Curl Virus (94.2% nucleotide sequence identity), which is known to infectSolanum lycopersicon, in Oman, whereas the 16S rRNA was identified from the sourceCandidatus Phytoplasma trifolii(99.04% nucleotide sequence identity), which is known to infect Helichrysum flowering plants in India. Subsequently, molecular computing research based on phylogenetic interweaves, putative recombination, amino acid selection, and genetic diversity were investigated, revealing divergent evolutionary patterns with significant variation and recombination events. The majority of the sequence variations observed in begomovirus and phytoplasma were caused via inter- and intra-specific recombination. These findings could be the firstin silicocombined infection analysis of ChiLCV andCa.P.trifoliiin a chilli crop in India, revealing the potential adaption and evolution of begomovirus and phytoplasma to a new geographic range and crop.

A dark-coloured thin film of cyanobacteria growing on the bottom of a submerged stone was isolated from Basantgarh village in Udhampur district, Jammu and Kashmir, India. The isolated strain (designated 19C-PST) was characterized using a polyphasic approach. The strain exhibited typical Nostoc -like morphology with a characteristic feature of having heterocytes in series. The 16S rRNA gene phylogeny placed the strain at a well-supported and distinct node. Notably, the recently described genus, Amazonocrinis , on the addition of more 16S rRNA gene sequences, reflected a critical split, which proved to be stable and well supported in all phylogenetic analyses of the 16S rRNA gene. Interestingly, Amazonocrinis nigriterrae CENA67T (type species of the genus) clustered together with our strain 19C-PST in the 16S rRNA gene phylogenetic analysis while the rest of the members of the genus Amazonocrinis were placed at a separate and distant node. This clearly indicated that strain 19C-PST is a member of Amazonocrinis sensu stricto. However, the results of phylogenetic analysis of ITS sequences only, in strains purported to belong to Amazonocrinis did not agree with the 16S rRNA gene results and placed our strain 19C-PST in a sister clade to three strains that have not yet been speciated, UHCC 0702, NIES-4103 and SA22, with A. nigriterrae falling into a separate clade. Further, folded secondary structures of the D1–D1′, V2, BoxB and V3 helices of strain 19C-PST were found to be significantly different from those of all the phylogenetically related taxa. The study revealed an interesting case where low taxon sampling and phylogenomic interpretations came across as points of attention in cyanobacterial taxonomy. Based on the morphological, phylogenetic, 16S–23S ITS secondary structure analyses, we describe our strain as Amazonocrinis malviyae sp. nov. in accordance with the International Code of Nomenclature for algae, fungi and plants. This work also illuminates the need for further research to resolve the taxonomic discrepancies among Amazonocrinis strains.

AbstractAkkermansiamuciniphila is a human intestinal tract bacterium that plays an important role in the mucus layer renewal. Several studies have demonstrated that it is a modulator for gut homeostasis and a probiotic for human health. The Akkermansia genus contains two species with standing in nomenclature but their genomic diversity remains unclear. In this study, eight new Akkermansia sp. strains were isolated from the human gut. Using the digital DNA-DNA hybridization (dDDH), average nucleotide identity (ANI) and core genome-based phylogenetic analysis applied to 104 A.muciniphila whole genomes sequences, strains were reclassified into three clusters. Cluster I groups A.muciniphila strains (including strain ATCC BAA-835T as type strain), whereas clusters II and III represent two new species. A member of cluster II, strain Marseille-P6666 differed from A.muciniphila strain ATCC BAA-835T and from A.glycaniphila strain PytT in its ability to grow in microaerophilic atmosphere up to 42 °C, to assimilate various carbon sources and to produce acids from a several compounds. The major fatty acids of strain Marseille-P6666 were 12-methyl-tetradecanoic and pentadecanoic acids. The DNA G + C content of strain Marseille-P6666 was 57.8%. On the basis of these properties, we propose the name A.massiliensis sp. nov. for members of cluster II, with strain Marseille-P6666T (= CSUR P6666 = CECT 30548) as type strain. We also propose the name “Candidatus Akkermansia timonensis” sp. nov. for the members of cluster III, which contains only uncultivated strains, strain Akk0196 being the type strain.

BackgroundHuanglongbing (HLB, yellow shoot disease) is a highly destructive citrus disease associated with a nonculturable bacterium, “Candidatus Liberibacter asiaticus” (CLas), which is transmitted by Asian citrus psyllid (ACP, Diaphorina citri). In Mexico, HLB was first reported in Tizimin, Yucatán, in 2009 and is now endemic in 351 municipalities of 25 states. Understanding the population diversity of CLas is critical for HLB management. Current CLas diversity research is exclusively based on analysis of the bacterial genome, which composed two regions, chromosome (> 1,000 genes) and prophage (about 40 genes).Methods and resultsIn this study, 40 CLas-infected ACP samples from 20 states in Mexico were collected. CLas was detected and confirmed by PCR assays. A prophage gene(terL)-based typing system (TTS) divided the Mexican CLas strains into two groups: Term-G including four strains from Yucatán and Chiapas, as well as strain psy62 from Florida, USA, and Term-A included all other 36 Mexican strains, as well as strain AHCA1 from California, USA. CLas diversity was further evaluated to include all chromosomal and prophage genes assisted by using machine learning (ML) tools to resolve multidimensional data handling issues. A Term-G strain (YTMX) and a Term-A strain (BCSMX) were sequenced and analyzed. The two Mexican genome sequences along with the CLas genome sequences available in GenBank were studied. An unsupervised ML was implemented through principal component analysis (PCA) on average nucleotide identities (ANIs) of CLas whole genome sequences; And a supervised ML was implemented through sparse partial least squares discriminant analysis (sPLS-DA) on single nucleotide polymorphisms (SNPs) of coding genes of CLas guided by the TTS. Two CLas Geno-groups, Geno-group 1 that extended Term-A and Geno-group 2 that extended Term-G, were established.ConclusionsThis study concluded that: 1) there were at least two different introductions of CLas into Mexico; 2) CLas strains between Mexico and USA are closely related; and 3) The two Geno-groups provide the basis for future CLas subspecies research.

’Candidatus Phytoplasma mali’, is a bacterial pathogen associated with the so-called apple proliferation disease in Malus × domestica. The pathogen manipulates its host with a set of effector proteins, among them SAP11CaPm, which shares similarity to SAP11AYWB from ’Candidatus Phytoplasma asteris’. SAP11AYWB interacts and destabilizes the class II CIN transcription factors of Arabidopsis thaliana, namely AtTCP4 and AtTCP13 as well as the class II CYC/TB1 transcription factor AtTCP18, also known as BRANCHED1 being an important factor for shoot branching. It has been shown that SAP11CaPm interacts with the Malus × domestica orthologues of AtTCP4 (MdTCP25) and AtTCP13 (MdTCP24), but an interaction with MdTCP16, the orthologue of AtTCP18, has never been proven. The aim of this study was to investigate this potential interaction and close a knowledge gap regarding the function of SAP11CaPm. A Yeast two-hybrid test and Bimolecular Fluorescence Complementation in planta revealed that SAP11CaPm interacts with MdTCP16. MdTCP16 is known to play a role in the control of the seasonal growth of perennial plants and an increase of MdTCP16 gene expression has been detected in apple leaves in autumn. In addition to this, MdTCP16 is highly expressed during phytoplasma infection. Binding of MdTCP16 by SAP11CaPm might lead to the induction of shoot proliferation and early bud break, both of which are characteristic symptoms of apple proliferation disease.