Microbiology (medical)

Phytoplasmas of the 16SrIII group are wide spread, and have a broad plant host range. Among these, ‘Candidatus phytoplasma pruni’ (‘Ca. P. pruni’; phytoplasmas of 16SrIII subgroup A) can cause serious diseases in Prunus species and ‘Ca. P. pruni’-related strains can infect other plant species, including grapevines. In this study, a new real-time PCR detection system was developed for ‘Ca. P. pruni’ using TaqMan chemistry. This test was designed to detect ‘Ca. P. pruni’, by amplifying the species-specific secY gene. In addition, a test to amplify the group-specific 16S rRNA gene region was also developed. The performances of both tests were evaluated. The test that amplifies the secY gene provided reliable and quick detection of ‘Ca. P. pruni’. Using the newly developed and validated test, ‘Ca. P. pruni’ was not found in any of the 434 field samples collected from different plants species grown in different regions of Slovenia.

Grapevine Bois noir (BN) is caused by ‘Candidatus Phytoplasma solani’ (‘Ca. P. solani’) and is one of the most important phytoplasma diseases in the Euro-Mediterranean viticultural areas. The epidemiology of BN can include grapevine as a plant host and is usually transmitted via sap-sucking insects that inhabit herbaceous host plants. Tracking the spread of ‘Ca. P. solani’ strains is of great help for the identification of plant reservoirs and insect vectors involved in local BN outbreaks. The molecular epidemiology of ‘Ca. P. solani’ is primarily based on sequence analysis of the tuf housekeeping gene (which encodes elongation factor Tu). In this study, molecular typing of tuf, through restriction fragment length polymorphism and sequencing, was carried out on grapevine samples from Iranian vineyards. According to the molecular characterization, three molecular types—tuf b1, tuf b5 and tuf b6—were found, with tuf b1 being the most prominent. These data provide further knowledge of tuf gene diversity and question the ecological role of such “minor” tuf types in Iranian vineyards, which have been detected only in grapevines.

Recently, we isolated two marine strains, F1T and F21T, which together with Kiritimatiella glycovorans L21-Fru-ABT are the only pure cultures of the class Kiritimatiellae within the phylum Verrucomicrobiota. Here, we present an in-depth genome-guided characterization of both isolates with emphasis on their exopolysaccharide synthesis. The strains only grew fermentatively on simple carbohydrates and sulfated polysaccharides. Strains F1T, F21T and K. glycovorans reduced elemental sulfur, ferric citrate and anthraquinone-2,6-disulfonate during anaerobic growth on sugars. Both strains produced exopolysaccharides during stationary phase, probably with intracellularly stored glycogen as energy and carbon source. Exopolysaccharides included N-sulfated polysaccharides probably containing hexosamines and thus resembling glycosaminoglycans. This implies that the isolates can both degrade and produce sulfated polysaccharides. Both strains encoded an unprecedently high number of glycoside hydrolase genes (422 and 388, respectively), including prevalent alpha-L-fucosidase genes, which may be necessary for degrading complex sulfated polysaccharides such as fucoidan. Strain F21T encoded three putative glycosaminoglycan sulfotransferases and a putative sulfate glycosaminoglycan biosynthesis gene cluster. Based on phylogenetic and chemotaxonomic analyses, we propose the taxa Pontiella desulfatans F1T gen. nov., sp. nov. and Pontiella sulfatireligans F21T sp. nov. as representatives of the Pontiellaceae fam. nov. within the class Kiritimatiellae.

Bat-associated bartonellae, including Bartonella mayotimonensis and Candidatus Bartonella rousetti, were recently identified as emerging and potential zoonotic agents, respectively. However, there is no report of bat-associated bartonellae in Zambia. Thus, we aimed to isolate and characterize Bartonella spp. from bats and bat flies captured in Zambia by culturing and PCR. Overall, Bartonella spp. were isolated from six out of 36 bats (16.7%), while Bartonella DNA was detected in nine out of 19 bat flies (47.3%). Subsequent characterization using a sequence of five different genes revealed that three isolates obtained from Egyptian fruit bats (Rousettus aegyptiacus) were Ca. B. rousetti. The isolates obtained from insectivorous bats (Macronycteris vittatus) were divided into two previously unclassified bat-associated bartonellae. A phylogenetic analysis of the six genotypes of Bartonella gltA sequences from nine pathogen-positive bat flies revealed that three genotypes belonged to the same clades as bat-associated bartonellae, including Ca. B. rousetti. The other three genotypes represented arthropod-associated bartonellae, which have previously been isolated only from ectoparasites. We demonstrated that Ca. B. rousetti is maintained between bats (R. aegyptiacus) and bat flies in Zambia. Continuous surveillance of Bartonella spp. in bats and serological surveys in humans in Africa are warranted to evaluate the public health importance of bat-associated bartonellae.

Bois noir (BN), associated with ‘Candidatus Phytoplasma solani’ (CaPsol), is the most widespread disease of the grapevine yellows complex worldwide. In this work, BN epidemiology was investigated in a case study vineyard where an unusual CaPsol strain, previously detected only in other host plants, was found to be prevalent in grapevine. Experimental activities included: symptom observation; sampling of symptomatic vines, Auchenorrhyncha specimens, and weeds; molecular detection and typing of CaPsol strains; statistical analyses for determining possible relationships between CaPsol relative concentration, strain type, and symptom severity. Among insects, Reptalus quinquecostatus was the most abundant and was found to be highly infected by CaPsol, while Hyalesthes obsoletus, the main CaPsol vector, was not caught. Moreover, R. quinquecostatus harbored CaPsol strains carrying uniquely the stamp sequence variant St10, also identified as prevalent in vines and in the majority of weeds, and all the secY variants identified in the vineyard. Statistical analyses revealed that CaPsol strains carrying the St10 variant are not associated with severe symptoms, suggesting their possible moderate virulence. Based on such evidence, a new BN epidemiological pattern related to these CaPsol strains and involving grapevine, R. quinquecostatus, and/or weeds is proposed. Furthermore, the possible presence of other players (vectors and weeds) involved in CaPsol transmission to grapevines was highlighted.