Publications

4470

The mitochondrial genomes of two vector psyllids of the ‘Candidatus Phytoplasma mali’, Cacopsylla picta and C. melanoneura, were sequenced using high-throughput sequencing on the Illumina platform. The main objective of the study was to describe their mitogenome and characterize their genetic variability and the potential changes in the context of the observed global warming. The four complete sequences for C. picta, 14,801 bp and 14,802 bp in length, two complete and one partial sequence for C. melanoneura, ranging from 14,879 bp to 14,881 bp in length, were obtained for the first time for these European apple psyllids. The detected intraspecies mtDNA identity was highly similar (99.85–99.98%), the identity’s similarity with other Cacopsylla species varied between 79.79 and 86.64%. The mitogenomes showed a typical mitochondrial DNA structure with 13 protein-coding genes, 2 rRNA genes and 22 tRNA genes; the presence of CGGA motif in the ND1-trnS2 junction was detected in both species. Phylogenetic analysis placed both species in close relationship with C. burckhardti within the Cacopsylla clade-I O group. The analysis of complete mitogenomes and of partial COI sequences of fifty-two Cacopsylla individuals showed a high homogeneity of genotypes over 15 years and among the different localities in the Czech Republic.

Symbiotic relationships are ubiquitous throughout the world’s oceans, yet for many marine organisms, including those in the high latitudes, little is understood about symbiotic associations and functional relationships. From a recently determined genome sequence of a filter-feeding basket star from Argentina, Gorgonocephalus chilensis, we discovered a novel Mycoplasma species with a 796Kb genome (CheckM completeness of 97.9%, G+C content = 30.1%). Similar to other Mycoplasma spp. within Mycoplasmatota, genomic analysis of the novel organism revealed reduced metabolic pathways including incomplete biosynthetic pathways, suggesting an obligate association with their basket star host. Results of 16S rRNA and multi-locus phylogenetic analyses revealed that this organism belonged to a recently characterized non-free-living lineage of Mycoplasma spp. specifically associated with marine invertebrate animals. Thus, the name “Candidatus Mycoplasma mahonii” is proposed for this novel species. Based on 16S rRNA PCR-screening, we found that Ca. M. mahonii also occurs in Gorgonocephalus eucnemis from the Northwest Pacific and other Gorgonocephalus chilensis from Argentinian waters. The level of sequence conservation within Ca. M. mahonii is considerable between widely disparate high-latitude Gorgonocephalus species, suggesting that oceanic dispersal of this microbe may be greater than excepted.

Ixodid ticks are responsible for the transmission of various intracellular bacteria, such as the Rickettsia species. Little Information is available about the genetic characterization and epidemiology of Rickettsia spp. The current study was designed to assess the tick species infesting various livestock hosts and the associated Rickettsia spp. in Pakistan. Ticks were collected from different livestock hosts (equids, cattle, buffaloes, sheep, goats, and camels); morphologically identified; and screened for the genetic characterization of Rickettsia spp. by the amplification of partial fragments of the gltA, ompA and ompB genes. Altogether, 707 ticks were collected from 373 infested hosts out of 575 observed hosts. The infested hosts comprised 105 cattle, 71 buffaloes, 70 sheep, 60 goats, 34 camels, and 33 equids. The overall occurrence of Rickettsia spp. was 7.6% (25/330) in the tested ticks. Rickettsia DNA was detected in Rhipicephalus haemaphysaloides (9/50, 18.0%), followed by Rhipicephalus turanicus (13/99, 13.1%), Haemaphysalis cornupunctata (1/18, 5.5%), and Rhipicephalus microplus (2/49, 4.1%); however, no rickettsial DNA was detected in Hyalomma anatolicum (71), Hyalomma dromedarii (35), and Haemaphysalis sulcata (8). Two Rickettsia agents were identified based on partial gltA, ompA, and ompB DNA sequences. The Rickettsia species detected in Rh. haemaphysaloides, Rh. turanicus, and Rh. microplus showed 99–100% identity with Rickettsia sp. and Candidatus Rickettsia shennongii, and in the phylogenetic trees clustered with the corresponding Rickettsia spp. The Rickettsia species detected in Rh. haemaphysaloides, Rh. turanicus, Rh. microplus, and Ha. cornupunctata showed 100% identity with R. massiliae, and in the phylogenetic trees it was clustered with the same species. Candidatus R. shennongii was characterized for the first time in Rh. haemaphysaloides, Rh. turanicus, and Rh. microplus. The presence of SFG Rickettsia spp., including the human pathogen R. massiliae, indicates a zoonotic risk in the study region, thus stressing the need for regular surveillance.

Bathyarchaeia are widespread in various anoxic ecosystems and are considered one of the most abundant microbial groups on the earth. There are only a few reports of laboratory cultivation of Bathyarchaeia, and none of the representatives of this class has been isolated in pure culture. Here, we report a sustainable cultivation of the Bathyarchaeia archaeon (strain M17CTs) enriched from anaerobic sediment of a coastal lake. The cells of strain M17CTs were small non-motile cocci, 0.4–0.7 μm in diameter. The cytoplasmic membrane was surrounded by an S-layer and covered with an outermost electron-dense sheath. Strain M17CTs is strictly anaerobic mesophile. It grows at 10–45°C (optimum 37°C), at pH 6.0–10.0 (optimum 8.0), and at NaCl concentrations of 0–60 g l−1 (optimum 20 g l−1). Growth occurred in the presence of methoxylated aromatic compounds (3,4-dimethoxybenzoate and vanillate) together with complex proteinaceous substrates. Dimethyl sulfoxide and nitrate stimulated growth. The phylogenomic analysis placed strain M17CTs to BIN-L-1 genus-level lineage from the BA1 family-level lineage and B26-1 order-level lineage (former subgroup-8) within the class Bathyarchaeia. The complete genome of strain M17CTs had a size of 2.15 Mb with a DNA G + C content of 38.1%. Based on phylogenomic position and phenotypic and genomic properties, we propose to assign strain M17CTs to a new species of a novel genus Bathyarchaeum tardum gen. nov., sp. nov. within the class Bathyarchaeia. This is the first sustainably cultivated representative of Bathyarchaeia. We propose under SeqCode the complete genome sequence of strain M17CTs (CP122380) as a nomenclatural type of Bathyarchaeum tardum, which should be considered as a type for the genus Bathyarchaeum, which is proposed as a type for the family Bathyarchaeaceae, order Bathyarchaeales, and of the class Bathyarchaeia.

AbstractBacteria of the SAR202 clade, within the phylum Chloroflexota, are ubiquitously distributed in the ocean but have not yet been cultivated in the lab. It has been proposed that ancient expansions of catabolic enzyme paralogs broadened the spectrum of organic compounds that SAR202 bacteria could oxidize, leading to transformations of the Earth’s carbon cycle. Here, we report the successful cultivation of SAR202 bacteria from surface seawater using dilution-to-extinction culturing. The growth of these strains is very slow (0.18–0.24 day−1) and is inhibited by exposure to light. The genomes, of ca. 3.08 Mbp, encode archaella (archaeal motility structures) and multiple sets of enzyme paralogs, including 80 genes coding for enolase superfamily enzymes and 44 genes encoding NAD(P)-dependent dehydrogenases. We propose that these enzyme paralogs participate in multiple parallel pathways for non-phosphorylative catabolism of sugars and sugar acids. Indeed, we demonstrate that SAR202 strains can utilize several substrates that are metabolized through the predicted pathways, such as sugars ʟ-fucose and ʟ-rhamnose, as well as their lactone and acid forms.

The pathogenicity of intracellular plant pathogenic bacteria is associated with the action of pathogenicity factors/effectors, but their physiological roles for most phytoplasma species, including ‘Candidiatus Phytoplasma solani’ are unknown. Six putative pathogenicity factors/effectors from six different strains of ‘Ca. P. solani’ were selected by bioinformatic analysis. The way in which they manipulate the host cellular machinery was elucidated by analyzing Nicotiana benthamiana leaves after Agrobacterium-mediated transient transformation with the pathogenicity factor/effector constructs using confocal microscopy, pull-down, and co-immunoprecipitation, and enzyme assays. Candidate pathogenicity factors/effectors were shown to modulate plant carbohydrate metabolism and the ascorbate–glutathione cycle and to induce autophagosomes. PoStoSP06, PoStoSP13, and PoStoSP28 were localized in the nucleus and cytosol. The most active effector in the processes studied was PoStoSP06. PoStoSP18 was associated with an increase in phosphoglucomutase activity, whereas PoStoSP28, previously annotated as an antigenic membrane protein StAMP, specifically interacted with phosphoglucomutase. PoStoSP04 induced only the ascorbate–glutathione cycle along with other pathogenicity factors/effectors. Candidate pathogenicity factors/effectors were involved in reprogramming host carbohydrate metabolism in favor of phytoplasma own growth and infection. They were specifically associated with three distinct metabolic pathways leading to fructose-6-phosphate as an input substrate for glycolysis. The possible significance of autophagosome induction by PoStoSP28 is discussed.

Based on genome-wide data, Massilia species belonging to the clade including Telluria mixta LMG 11547T should be entirely transferred to the genus Telluria owing to the nomenclatural priority of the type species Telluria mixta . This results in the transfer of 35 Massilia species to the genus Telluria . The presented data also supports the creation of two new genera since peripherally branching Massilia species are distinct from Telluria and other related genera. It is proposed that 13 Massilia species are transferred to Mokoshia gen. nov. with the type species designated Mokoshia eurypsychrophila comb. nov. The species Massilia arenosa is proposed to belong to the genus Zemynaea gen. nov. as the type species Zemynaea arenosa comb. nov. The genome-wide analysis was well supported by canonical ordination analysis of Enzyme Commission (EC) codes annotated from genomes via pannzer2. This new approach was performed to assess the conclusions of the genome-based data and reduce possible ambiguity in the taxonomic decision making. Cross-validation of EC code data compared within canonical plots validated the reclassifications and correctly visualized the expected genus-level taxonomic relationships. The approach is complementary to genome-wide methodology and could be used for testing sequence alignment based data across genetically related genera. In addition to the proposed broader reclassifications, invalidly described species ‘Massilia antibiotica’, ‘Massilia aromaticivorans’, ‘Massilia cellulosiltytica’ and ‘ Massilia humi ’ are described as Telluria antibiotica sp. nov., Telluria aromaticivorans sp. nov., Telluria cellulosilytica sp. nov. and Pseudoduganella humi sp. nov., respectively. In addition, Telluria chitinolytica is reclassified as Pseudoduganella chitinolytica comb. nov. The use of combined genome-wide and annotation descriptors compared using canonical ordination clarifies the taxonomy of Telluria and its sibling genera and provides another way to evaluate complex taxonomic data.

Abstract Autophagy functions in plant host immunity responses to pathogen infection. The molecular mechanisms and functions used by the citrus Huanglongbing (HLB)-associated intracellular bacterium ‘Candidatus Liberibacter asiaticus’ (CLas) to manipulate autophagy are unknown. We identified a CLas effector, SDE4405 (CLIBASIA_04405), which contributes to HLB progression. ‘Wanjincheng’ orange (Citrus sinensis) transgenic plants expressing SDE4405 promotes CLas proliferation and symptom expression via suppressing host immunity responses. SDE4405 interacts with the ATG8-family of proteins (ATG8s), and their interactions activate autophagy in Nicotiana benthamiana. The occurrence of autophagy is also significantly enhanced in SDE4405-transgenic citrus plants. Interrupting NbATG8s-SDE4405 interaction by silencing of NbATG8c reduces Pseudomonas syringae pv. tomato strain DC3000ΔhopQ1-1 (Pst DC3000ΔhopQ1-1) proliferation in N. benthamiana, and transient overexpression of CsATG8c and SDE4405 in citrus promotes Xanthomonas citri subsp. citri (Xcc) multiplication, suggesting that SDE4405-ATG8s interaction negatively regulates plant defense. These results demonstrate the role of the CLas effector protein in manipulating autophagy, and provide new molecular insights into the interaction between CLas and citrus hosts.