Publications

3989

ABSTRACTBiogas reactors operating at elevated ammonia levels are commonly susceptible to process disturbances, further augmented at thermophilic temperatures. The major cause is assumed to be linked to inhibition followed by an imbalance between different functional microbial groups, centred around the last two steps of the anaerobic digestion, involving acetogens, syntrophic acetate oxidisers (SAOB) and methanogens. Acetogens are key contributors to reactor efficiency, acting as the crucial link between the hydrolysis and fermentation steps and the final methanogenesis step. Their major product is acetate, at high ammonia levels further converted by SAOB and hydrogenotrophic methanogens to biogas. Even though these functionally different processes are well recognised, less is known about the responsible organism at elevated temperature and ammonia conditions. The main aim of this study was to garner insights into the penultimate stages in three thermophilic reactors (52°C) operated under high ammonia levels (FAN 0.7–1.0 g/L; TAN 3.6–4.4 g/L). The primary objective was to identify potential acetogens and SAOBs. Metagenomic data from the three reactors were analysed for the reductive acetyl‐CoA pathway (Wood–Ljungdahl Pathway) and glycine synthase reductase pathway. The results revealed a lack of true acetogens but uncovered three potential SAOB candidates that harbour the WLP, ‘Candidatus Thermodarwinisyntropha acetovorans’, ‘Candidatus Thermosyntrophaceticus schinkii’, ‘Candidatus Thermotepidanaerobacter aceticum’, and a potential lipid‐degrader ‘Candidatus Thermosyntrophomonas ammoiaca’.

AbstractBACKGROUNDHuanglongbing (HLB) is a systemic disease of citrus caused by the bacterial pathogen Candidatus Liberibacter asiaticus (CLas) that limits citrus production worldwide. CLas is an obligate bacterial pathogen that multiplies in citrus trees and in the insect vector, the Asian citrus psyllid (ACP), Diaphorina citri Kuwayama. There is no cure for HLB currently and broad‐spectrum antibiotics represent one possible therapeutic against disease symptoms. Single‐stranded nucleic acid analogs, 2′‐deoxy‐2’‐Fluoro‐β‐D‐arabinonucleic antisense oligonucleotides (FANA ASOs), can modulate gene expression by enzymatic degradation or steric blocking of an RNA target. FANA ASOs recognize and bind to specific RNA forms, including mRNA, miRNA and long noncoding RNA, through complementary base‐pairing.RESULTSInjection of oxytetracycline (OTC) into mature citrus trees with HLB ameliorated symptoms of disease, increased fruit yield and quality of juice as compared with that produced by noninjected controls. Injection of trees with FANA ASOs reduced CLas abundance but did not improve fruit yield and quality above control levels at the injection dosage tested. Reduced pathogen relative abundance following OTC injection was coincident with lower CLas acquisition and inoculation by laboratory deployed and wild‐type D. citri collected from the field, respectively. However, injections of FANA ASOs did not consistently reduce CLas transmission as compared with noninjected controls.CONCLUSIONSTrunk injection of OTC may be useful in management of HLB by reducing CLas infection in trees and disrupting transmission. However, more research is needed to verify the potential usefulness of FANA ASOs. Optimizing FANA ASO dosage in trees and exploring the potential of multiplex FANA ASOs that simultaneously target multiple mRNAs could possibly enhance their efficacy against CLas. © 2024 Society of Chemical Industry.

The occurrence of ‘Candidatus Liberibacter’ spp. and ‘Ca. Phytoplasma’ spp. associated with blotchy mottle symptoms poses challenges to huanglongbing (HLB) diagnosis using molecular techniques. The ability to detect multiple targets simultaneously and specifically is a key aspect met by quantitative PCR (qPCR). A set of primers and hydrolysis probes useful in either single or multiplex reactions for the detection and quantification of HLB-associated bacteria were developed. Sequences from conserved genes of the ribosomal proteins for Liberibacter and phytoplasma circumvent the lack of specificity and cross-reactivity problems related to 16Sr DNA gene amplification, allowing precise and specific detection of HLB-associated bacteria in citrus and in the Liberibacter vector, Diaphorina citri. The triplex reaction exhibited high quality and precision as a robust tool for quantifying ‘Ca. L. asiaticus’ (CLas), ‘Ca. L. americanus’ (CLam), and 16Sr IX phytoplasma. Triplex qPCR showed consistent results and comparable sensitivity to the ribonuclease reductase test, although quantification cycle (Cq) values were higher when compared with 16SrDNA qPCR. Detection tests using field samples indicate that the qPCR triplex can identify HLB-associated bacteria in samples with varying levels of symptoms, ranging from typical to asymptomatic. Assessment of field samples from growers indicated more than 78.6% had Cq lower than 35.0, below the cutoff established for qPCR reactions used in this work. qPCR triplex is a safe, specific, and sufficiently sensitive technique for detecting CLas, CLam, and 16Sr IX phytoplasma simultaneously, in both citrus and D. citri samples. Its application is of importance in assisting growers in making decisions for HLB management.

Opinion 131 addresses a Request for an Opinion asking the Judicial Commission to conserve the genus name Proteus Hauser 1885 (Approved Lists 1980) over its earlier homonym, the protozoan genus name Proteus Müller 1786. The Judicial Commission agrees that the later homonym is illegitimate and that the replacement of the prokaryotic name Proteus would be undesirable. It is also concluded that Proteus Müller 1786 is an objectively invalid name under the International Code of Zoological Nomenclature. Judicial Opinions 9, 12 and 130 serve as precedents for the conservation of Proteus Hauser 1885 (Approved Lists 1980) over Proteus Müller 1786. This action is taken here and makes the prokaryotic name Proteus legitimate.