Publications

4467

Huanglongbing is a highly destructive citrus disease associated with “Candidatus Liberibacter asiaticus” (Las), a phloem−limited and non-culturable bacterium, naturally transmitted by the psyllid Diaphorina citri. Although diverse approaches have been used to understand the molecular mechanisms involved in the pathogen–host interaction, such approaches have focused on already infected and/or symptomatic plants, missing early events in the initial days post-inoculation. This study aimed to identify the time course of Las multiplication and whole-plant colonization immediately following inoculation by infected psyllids feeding for 2 days. Thus, the experimental approach was to track Las titers after psyllid inoculation in new shoots (NS) of Citrus × sinensis (susceptible), Murraya paniculata (partially resistant), and Bergera koenigii (fully resistant). Soon after psyllid removal, Las titers dropped until the 10–12th days in all three species. Following this, Las titers increased exponentially only in C. × sinensis and M. paniculata, indicating active bacterial multiplication. In C. × sinensis, Las reached a stationary phase at ∼5 log Las cells/g of tissue from the 40th day onward, while in M. paniculata, Las increased at a lower rate of up to ∼3 log Las cells/g of tissue between the 40th and 60th days, decreasing gradually thereafter and becoming undetectable from the 160th day onward. In B. koenigii, Las titers decreased from the start and remained undetectable. In C. × sinensis, an average of 2.6 log of Las cells/g of tissue was necessary for Las to move out of 50% of the NS in 23.6 days and to colonize the rest of the plant, causing a successful infection. Conversely, the probability of Las moving out of the NS remained below 50% in M. paniculata and zero in B. koenigii. To our knowledge, this is the first study on Las dynamics and whole-plant colonization during the earliest stages of infection. Identification of critical time-points for either successful multiplication or Las resistance may help to elucidate initial events of Las–host interactions that may be missed due to longer sampling intervals and at later stages of infection.

A novel Borrelia species, Candidatus Borrelia javanense, was found in ectoparasite ticks, Amblyomma javanense, from Manis javanica pangolins seized in anti-smuggling operations in southern China. Overall, 12 tick samples in 227 (overall prevalence 5.3%) were positive for Candidatus B. javanense, 9 (5.1%) in 176 males, and 3 (5.9%) in 51 females. The phylogenetic analysis, based on the 16S rRNA gene and the flagellin gene sequences of the Borrelia sp., exhibited strong evidence that Candidatus B. javanense did not belong to the Lyme disease Borrelia group and the relapsing fever Borrelia group but another lineage of Borrelia. The discovery of the novel Borrelia species suggests that A. javanense may be the transmit vector, and the M. javanica pangolins should be considered a possible origin reservoir in the natural circulation of these new pathogens. To our knowledge, this is the first identification of a novel Borrelia species agent in A. javanense from pangolins. Whether the novel agent is pathogenic to humans is unknown and needs further research.

Citrus Huanglongbing (HLB) is the most devastating disease of citrus, presumably caused by “Candidatus Liberibacter asiaticus” (CaLas). Although transcriptomic profiling of HLB-affected citrus plants has been studied extensively, the initial steps in pathogenesis have not been fully understood. In this study, RNA sequencing (RNA-seq) was used to compare very early transcriptional changes in the response of Valencia sweet orange (VAL) to CaLas after being fed by the vector, Diaphorina citri (Asian citrus psyllid, or ACP). The results suggest the existence of a delayed defense reaction against the infective vector in VAL, while the attack by the healthy vector prompted immediate and substantial transcriptomic changes that led to the rapid erection of active defenses. Moreover, in the presence of CaLas-infected psyllids, several downregulated differentially expressed genes (DEGs) were identified on the pathways, such as signaling, transcription factor, hormone, defense, and photosynthesis-related pathways at 1 day post-infestation (dpi). Surprisingly, a burst of DEGs (6,055) was detected at 5 dpi, including both upregulated and downregulated DEGs on the defense-related and secondary metabolic pathways, and severely downregulated DEGs on the photosynthesis-related pathways. Very interestingly, a significant number of those downregulated DEGs required ATP binding for the activation of phosphate as substrate; meanwhile, abundant highly upregulated DEGs were detected on the ATP biosynthetic and glycolytic pathways. These findings highlight the energy requirement of CaLas virulence processes. The emerging picture is that CaLas not only employs virulence strategies to subvert the host cell immunity, but the fast-replicating CaLas also actively rewires host cellular metabolic pathways to obtain the necessary energy and molecular building blocks to support virulence and the replication process. Taken together, the very early response of citrus to the CaLas, vectored by infective ACP, was evaluated for the first time, thus allowing the changes in gene expression relating to the primary mechanisms of susceptibility and host–pathogen interactions to be studied, and without the secondary effects caused by the development of complex whole plant symptoms.

In Iran, polyphasic studies of unicellular cyanobacteria are still scarce, with more emphasis being placed on filamentous cyanobacteria in paddy fields and fresh water regions. In an effort to increase the knowledge of the diversity of unicellular cyanobacteria from paddy fields in Iran, we have isolated and characterized a new unicellular cyanobacterium strain. The strain was studied using a polyphasic approach based on morphological, ecological and phylogenetic analyses of the 16S–23S ITS rRNA gene region. Complementarily, we have searched for the presence of cyanotoxin genes and analysed the pigment content of the strain. Results showed that the strain was morphologically indistinguishable from the genus Chroococcus , but phylogenetic analyses based on the Bayesian inference and maximum-likelihood methods placed the strain in a separated monophyletic and highly supported (0.99/98, posterior probability/maximum-likelihood) genus-level cluster, distant from Chroococcus sensu stricto and with Chalicogloea cavernicola as sister taxa. The calculated p-distance for the 16S rRNA gene also reinforced the presence of a new genus, by showing 92 % similarity to C. cavernicola . The D1–D1′, Box-B and V3 ITS secondary structures showed the uniqueness of this strain, as it shared no similar pattern with closest genera within the Chroococcales. For all these reasons, and in accordance with the International Code of Nomenclature for Algae, Fungi and Plants, we here proposed the description of a new genus with the name Alborzia gen. nov. along with the description of a new species, Alborzia kermanshahica sp. nov. (holotype: CCC1399-a; reference strains CCC1399-b; MCC 4116).

Abstract 'Ca. Phytoplasma pini' is a member of phytoplasma 16S rRNA gene RFLP group XXI, subgroup XXI-A. It has been identified in a number of European countries, including Germany, Poland, Lithuania, Spain, Czech Republic and Croatia. Outside of Europe it has been identified in China and Mozambique. A related strain has also been identified in Maryland, USA. In the USA, favourable climatic conditions and wide availability of potential host plants of the phytoplasma, suggest that the potential for spread of 'Ca. Phytoplasma pini' could be significant. Host plants include Pinus sylvestris, P. halepensis, P. mugo, P. banksiana, P. nigra, P. tabuliformis, Abies procera and Tsuga canadensis. Symptoms include the formation of ball-like growths containing dwarfed needles, yellowed or reddish needles and the loss of needles. It is transmitted by insect vectors that are currently unknown.

Abstract Phytoplasmas are wall-less parasitic bacteria living exclusively in plant phloem as consequence of transmission by sap-sucking insect vectors (Lee et al., 2000); they have been associated with several hundred plant diseases. 'Candidatus Phytoplasma phoenicium' (CaPphoe), subgroup 16SrIX-B, is the aetiological agent of almond witches'-broom (AlmWB), a severe disease affecting almond, peach and nectarine trees in Lebanon and Iran. The first epidemics of AlmWB occurred in almond trees in Lebanon in the early 1990s and in Iran in 1995. In Lebanon, the disease rapidly spread from coastal to high mountainous areas, killing almost 150,000 trees over a period of 15 years. CaPphoe was first added to the EPPO Alert List in 2001 and removed from the list in 2006. The more recent rapid spread of CaPphoe in peach and nectarine orchards and in other plant hosts, along with the identification of efficient insect vectors, increased the alarm about the risk it poses for stone fruit production in the Middle East and in all the countries of the Mediterranean basin. Thus it was re-inserted in the EPPO Alert List in 2015.

Abstract.We tried to determine the epidemiology and species of human dirofilariasis observed at two tertiary care hospitals in Kerala. We searched the hospital database to identify cases of dirofilariosis from January 2005 to March 2020. Along with human isolates, one dog Dirofilaria isolate was also subjected to PCR and sequencing of pan filarial primers cytochrome oxidase subunits 1 and 12S rDNA. We documented 78 cases of human dirofilariosis. The orbit, eyelid, and conjunctiva were the most commonly affected sites. Molecular characterization identified one dog and five human isolates as Candidatus Dirofilaria Hongkongensis. A rare case of subconjunctival infestation by B. malayi was also documented. Human dirofilariosis is a public health problem in the state of Kerala in India, and it is mostly caused by Candidatus Dirofilaria Hongkongensis. We propose that all diroifilaria isolates are subjected to sequencing for identification.

Abstract Phytoplasmas are cell-wall-less plant pathogenic bacteria of the class Mollicutes, which inhabit the phloem sieve tubes of plants and have been associated with several hundred diseases affecting economically important crops. Over the past few decades 'Candidatus Phytoplasma solani', belonging to the 16SrXII-A ribosomal subgroup, has been found to cause a range of plant diseases in different agro-ecosystems in many countries in Europe and the eastern Mediterranean area and a number of others all over the world. It is thought likely that it has always been present, at least in its European range, but has only been noticed in recent years. Diseases caused include bois noir in grapevines, stolbur in tomatoes, potatoes and other wild and cultivated plants, maize redness, lavender decline, and yellowing, reddening, decline, dwarfism, leaf malformation and degeneration diseases of other plants. 'Ca. P. solani' is usually transmitted from plant to plant by the polyphagous insect vector Hyalesthes obsoletus (Cixiidae) which, although it can complete its life cycle on only a small number of plant species, feeds on a much wider range. Recent studies have demonstrated the presence of additional insect vectors of this phytoplasma in Europe, such as Reptalus panzeri in Serbia, possibly R. quinquecostatus in Serbia and France, and Anaceratagallia ribauti in Austria. This scenario highlights the extreme complexity of the ecology of both 'Ca. Phytoplasma solani' and its insect vectors, underlying the difficulty in studying the epidemiology of diseases associated with this pathogen and in developing efficient control strategies. 'Ca. Phytoplasma solani' is also transmitted by parasitic plants and by grafting and vegetative propagation of infected host plants; it can be spread when host plants are transported by people. In the European Union it is listed as a harmful organism necessitating restrictions on the import of plants in the family Solanaceae.

Abstract The reference strain of 'Ca. Phytoplasma trifolii' is the causative agent of clover proliferation (CP) disease of alsike clover (Trifolium hybridum). The CP disease was first reported in Canada in the early 1960s when the aetiological agent was mistakenly presumed to be a yellows-type virus (Chiykowski, 1965). Subsequent investigations revealed that the disease was associated with infection by a mycoplasma-like organism (Chen and Hiruki, 1975>; Hiruki and Chen, 1984), now termed phytoplasma, strain CPR (Hiruki and Wang, 2004). Later, phytoplasmas of the same lineage (subgroup 16SrVI-A) were found in the USA, Mexico, and many countries in Europe and Asia, causing diseases in diverse leguminous and vegetable crops, responsible for significant yield losses and quality reductions. Phytoplasmas of the same lineage also caused disease in elm trees in the USA. Phytoplasmas of closely-related lineages (various subgroups of group 16SrVI) also have wide distributions around the world.

Abstract Phytoplasmas are wall-less, phloem-limited unculturable bacteria that are naturally spread by sap-sucking insects. 'Candidatus Phytoplasma australiense', subgroup 16SrXII-B, is associated with a wide range of diseases in Australia and New Zealand. Important commercial crop hosts of 'Ca. Phytoplasma australiense' include grapevine, papaya and strawberry. This phytoplasma is associated with rapid death of its papaya and cabbage tree hosts. In New Zealand, the insect vectors have been confirmed to be the endemic Cixiid planthoppers, Zeoliarus atkinsoni and Z. oppositus, while in Australia no vector has yet been determined, although the leafhopper, Orosius argentatus, has been implicated. Long distance spread of the phytoplasma is possible through infected vegetative propagating material. 'Ca. Phytoplasma australiense' is on the A1 list of regulated organisms for Canada and Bahrain, and is listed as a quarantine pest for the USA.