Publications

4369

Anaerobic methanotrophic (ANME) archaea can drive anaerobic oxidation of methane (AOM) using solid iron or manganese oxides as the electron acceptors, hypothetically via direct extracellular electron transfer (EET). This study investigated the response of Candidatus “Methanoperedens nitroreducens TS” (type strain), an ANME archaeon previously characterized to perform nitrate-dependent AOM, to an Fe(III)-amended condition over a prolonged period. Simultaneous consumption of methane and production of dissolved Fe(II) were observed for more than 500 days in the presence of Ca. “M. nitroreducens TS,” indicating that this archaeon can carry out Fe(III)-dependent AOM for a long period. Ca. “M. nitroreducens TS” possesses multiple multiheme c-type cytochromes (MHCs), suggesting that it may have the capability to reduce Fe(III) via EET. Intriguingly, most of these MHCs are orthologous to those identified in Candidatus “Methanoperedens ferrireducens,” an Fe(III)-reducing ANME archaeon. In contrast, the population of Ca. “M. nitroreducens TS” declined and was eventually replaced by Ca. “M. ferrireducens,” implying niche differentiation between these two ANME archaea in the environment.

Anaerobic methanotrophic (ANME) archaea have recently been reported to be capable of using insoluble extracellular electron acceptors via extracellular electron transfer (EET). In this study, we investigated EET by a microbial community dominated by “Candidatus Methanoperedens” archaea at the anode of a bioelectrochemical system (BES) poised at 0 V vs. standard hydrogen electrode (SHE), in this way measuring current as a direct proxy of EET by this community. After inoculation of the BES, the maximum current density was 274 mA m–2 (stable current up to 39 mA m–2). Concomitant conversion of 13CH4 into 13CO2 demonstrated that current production was methane-dependent, with 38% of the current attributed directly to methane supply. Based on the current production and methane uptake in a closed system, the Coulombic efficiency was about 17%. Polarization curves demonstrated that the current was limited by microbial activity at potentials above 0 V. The metatranscriptome of the inoculum was mined for the expression of c-type cytochromes potentially used for EET, which led to the identification of several multiheme c-type cytochrome-encoding genes among the most abundant transcripts in “Ca. Methanoperedens.” Our study provides strong indications of EET in ANME archaea and describes a system in which ANME-mediated EET can be investigated under laboratory conditions, which provides new research opportunities for mechanistic studies and possibly the generation of axenic ANME cultures.

Amaç: Bu çalışmada Elma Çoklu Sürgün Fitoplazma Hastalığı (EÇSF) (Candidatus Pyhtoplasma mali)’nın biyolojik mücadelesinde endofitik fungusların etkinliğinin belirlenmesi amaçlanmıştır. Böylece pratikte kimyasal mücadele olanağı olmayan hastalık etmenine karşı farklı kontrol stratejileri geliştirilebilme olanağının sağlanması hedeflenmiştir. Yöntem ve Bulgular: EÇSF ve endofit fungus izolasyonları için toplam 50 farklı bahçe gezilerek fitoplazma testleri için 360 ağaçtan, endofit funguslar için ise hem sağlıklı hem de fitoplazma ile enfekteli 198 ağaçtan bitki örneği toplanmıştır. Fitoplazma açısından testlenen 360 örnekten 29 tanesi ‘Ca. P. mali’ ile enfekteli bulunmuştur. ‘Ca. P. mali’ ile enfekteli ve sağlıklı elma ağaçlarından endofit fungus türleri izole edilmiş, bu fungusların morfolojik ve moleküler tanısı ile birlikte patojenisite çalışmaları yapılmıştır. Sağlıklı elma ağaçlarından en yaygın olarak izole edilen endofit funguslardan Epicoccum nigrum, Aureobasidium pullulans, Ulocladium consortiale’nin EÇSF hastalığına karşı etkinlikleri incelenmiştir. Her bir fungus izolatının EÇSF hastalık yoğunluğunu farklı düzeylerde etkilediği, en yüksek etkinin A. pullulans’a ait olduğu ve EÇSF hastalık yoğunluğunu uygulama sonrası %34,18 etki değeri ile azalttığı belirlenmiştir. Genel Yorum: EÇSF hastalığının etkili bir kimyasal mücadelesinin bulunmaması ve etmenin karantina listesinde yer alması nedeniyle olası biyolojik mücadele uygulamaları oldukça önemlidir. EÇSF ile enfekteli ağaçlara inokule edilen 3 farklı endofit fungus türünden Aureobasidium pullulans’ın fitoplazma yoğunluğunu diğer 2 etmene göre daha fazla azaltmasından dolayı A. pullulans hastalıkla mücadelede kullanılabilecek biyolojik mücadele ajanı olabilir. Çalışmanın Önemi ve Etkisi: Bu çalışma ile biyolojik mücadele ajanı olan endofit fungusların elmalarda EÇSF hastalığı üzerine etkileri Türkiye’de ilk kez araştırılmış ve hastalığın kontrol stratejilerinin oluşturulmasında önemli ön bulgular elde edilmiştir.

AbstractGeothermal environments, including terrestrial hot springs and deep-sea hydrothermal sediments, often contain many poorly understood lineages of archaea. Here, we recovered ten metagenome-assembled genomes (MAGs) from geothermal sediments and propose that they constitute a new archaeal class within the TACK superphylum, “Candidatus Culexarchaeia”, named after the Culex Basin in Yellowstone National Park. Culexarchaeia harbor distinct sets of proteins involved in key cellular processes that are either phylogenetically divergent or are absent from other closely related TACK lineages, with a particular divergence in cell division and cytoskeletal proteins. Metabolic reconstruction revealed that Culexarchaeia have the capacity to metabolize a wide variety of organic and inorganic substrates. Notably, Culexarchaeia encode a unique modular, membrane associated, and energy conserving [NiFe]-hydrogenase complex that potentially interacts with heterodisulfide reductase (Hdr) subunits. Comparison of this [NiFe]-hydrogenase complex with similar complexes from other archaea suggests that interactions between membrane associated [NiFe]-hydrogenases and Hdr may be more widespread than previously appreciated in both methanogenic and non-methanogenic lifestyles. The analysis of Culexarchaeia further expands our understanding of the phylogenetic and functional diversity of lineages within the TACK superphylum and the ecology, physiology, and evolution of these organisms in extreme environments.

Abstract‘Candidatus Phytoplasma ulmi’, associated with elm yellows, shoot proliferation and dieback of elm (Ulmus) species trees was reported in United States of America and in many European countries. Until now its presence in elm trees in Poland has not been detected. In 2017–2018, during visual inspection of elm trees grown in four areas of southern Silesia Province leaf yellowing, shoot proliferation, phloem necrosis and dieback of branches were observed on European field elm trees grown in nature reserve close to the border with the Czech Republic. Phloem tissue from shoots collected from six symptomatic and twenty asymptomatic trees was used for extraction nucleic acids. Nested PCR with primer pairs amplifying 16Sr RNA sequences of phytoplasmas resulted in specific products for samples from six symptomatic trees however the restriction fragment length polymorphism (RFLP) and phylogenetic analyses of the sequenced products did not allow to clearly classify newly detected isolates within the elm yellows group (16SrV). The isolates found in Poland clustered on the boundary of the 16SrV‐A and 16SrV‐C ribosomal subgroups. Sequence analyses of less conservative genes rpl22‐ rps3 and secY confirmed the affiliation of the phytoplasma isolates found in Poland to ‘Candidatus Phytoplasma ulmi’; however, the significant nucleotide changes in signature sequences for all three genes were shown compared with EY1T reference strain. Surprisingly, the newly detected isolates shared a common origin and were phylogenetically closer to EY1‐SRB and EY10‐SRB strains from Serbia as well as NK16 strain from Croatia strains than to the EYCZ1 strain from the Czech Republic. This is the first report on detection and molecular characterization of ‘Candidatus Phytoplasma ulmi’ in Poland based on sequence analysis of three genetic locus.

AbstractPear decline, induced by the phytoplasma 'CandidatusPhytoplasma pyri', transmitted by pear psyllids, is one of the most devastating diseases onPyrus communisin Europe and North America. Investigations of pear psyllids in 4 pear orchards in lower Austria showed the presence ofCacopsylla pyri,C. pyricolaandC. pyrisugaat all locations. PCR analyses revealed overall phytoplasma infection rates forC. pyriof 5.4%, forC. pyricola,of 4.6%, forC. pyrisugaremigrants of 9.6% and forC. pyrisugaemigrants of 0%. The rates of PCR-positiveC. pyriandC. pyricolaindividuals varied greatly in the course of the year, and the highest infection rates were observed in late summer, autumn and in late winter. In transmission experiments with healthy pear seedlings, winterform individuals ofC. pyri and C. pyricolatransmitted the pathogen to 19.2% (5 out of 26) and 4.8% (2 out of 41) of the test plants, respectively. The vectoring ability ofC. pyrisugawas experimentally proven for the first time, and in transmission experiments with remigrants, 9.5% (2 out of 21) of the pear seedlings were infected. Our data indicate a significant risk of pathogen transmission in pear orchards during the greater part of the year, especially in late winter, early spring and autumn. Multilocus sequence analysis by aid of the genesaceFandimpallowed the discrimination between 15 phytoplasma types. Three so far undescribedaceFgenotypes and four undescribedimpgenotypes were identified.

Summary Complete genomes can be recovered from metagenomes by assembling and binning DNA sequences into metagenome assembled genomes (MAGs). Yet, the presence of microdiversity can hamper the assembly and binning processes, possibly yielding chimeric, highly fragmented and incomplete genomes. Here, the metagenomes of four samples of aerobic granular sludge bioreactors containing Candidatus ( Ca .) Accumulibacter, a phosphate‐accumulating organism of interest for wastewater treatment, were sequenced with both PacBio and Illumina. Different strategies of genome assembly and binning were investigated, including published protocols and a binning procedure adapted to the binning of long contigs (MuLoBiSC). Multiple criteria were considered to select the best strategy for Ca . Accumulibacter, whose multiple strains in every sample represent a challenging microdiversity. In this case, the best strategy relies on long‐read only assembly and a custom binning procedure including MuLoBiSC in metaWRAP. Several high‐quality Ca . Accumulibacter MAGs, including a novel species, were obtained independently from different samples. Comparative genomic analysis showed that MAGs retrieved in different samples harbour genomic rearrangements in addition to accumulation of point mutations. The microdiversity of Ca . Accumulibacter, likely driven by mobile genetic elements, causes major difficulties in recovering MAGs, but it is also a hallmark of the panmictic lifestyle of these bacteria.