Methylated chalcones are required for rhizobial nod gene induction in the Medicago truncatula rhizosphere.

Legume nodulation requires the detection of flavonoids in the rhizosphere by rhizobia to activate their production of Nod factor countersignals. Here we investigated the flavonoids involved in nodulation of Medicago truncatula. We biochemically characterized five flavonoid-O-methyltransferases (OMTs) and a lux-based nod gene reporter was used to investigate the response of Sinorhizobium medicae NodD1 to various flavonoids. We found that chalcone-OMT 1 (ChOMT1) and ChOMT3, but not OMT2, 4, and 5, were able to produce 4,4'-dihydroxy-2'-methoxychalcone (DHMC). The bioreporter responded most strongly to DHMC, while isoflavones important for nodulation of soybean (Glycine max) showed no activity. Mutant analysis revealed that loss of ChOMT1 strongly reduced DHMC levels. Furthermore, chomt1 and omt2 showed strongly reduced bioreporter luminescence in their rhizospheres. In addition, loss of both ChOMT1 and ChOMT3 reduced nodulation, and this phenotype was strengthened by the further loss of OMT2. We conclude that: the loss of ChOMT1 greatly reduces root DHMC levels; ChOMT1 or OMT2 are important for nod gene activation in the rhizosphere; and ChOMT1/3 and OMT2 promote nodulation. Our findings suggest a degree of exclusivity in the flavonoids used for nodulation in M. truncatula compared to soybean, supporting a role for flavonoids in rhizobial host range.

Anthocyanin pigmentation as a quantitative visual marker for arbuscular mycorrhizal fungal colonization of Medicago truncatula roots.

Visualization of root colonization by arbuscular mycorrhizal fungi (AMF) is the most elementary experiment in the field of mycorrhizal symbiosis. The most widely used approach for evaluating levels of AMF colonization is staining with trypan blue or ink, which is scored using the time-consuming grid intersection method. Here we demonstrate the use of an anthocyanin-based visual marker system for visualizing AMF colonization of Medicago truncatula roots. Expression of MtLAP1, a transcription factor which regulates the production of anthocyanins, from the AMF-induced Kunitz Protease Inhibitor 106 promoter, allowed the visualization of arbuscules in live plant tissues without microscopy or staining. This marker system allowed straightforward qualitative evaluation of the ram1, vpy and dmi3 AMF phenotypes using Agrobacterium rhizogenes hairy-root transformation. For the strigolactone biosynthesis mutant carotenoid cleavage dioxygenase 8a and a novel mutant scooby, which show quantitative AMF symbiotic phenotypes, the amount of anthocyanins in the roots estimated by spectrophotometry correlated very well with colonization levels estimated by staining and scoring using the grid intersection method. The LAP1-based marker system therefore provides a highly efficient approach for mutant screening and monitoring of AMF colonization in live tissues by eye, or for quantitative assessment using a simple and quick photometric assay.

Three Common Symbiotic ABC Subfamily B Transporters in Medicago truncatula Are Regulated by a NIN-Independent Branch of the Symbiosis Signaling Pathway.

Several ATP-binding cassette (ABC) transporters involved in the arbuscular mycorrhizal symbiosis and nodulation have been identified. We describe three previously unreported ABC subfamily B transporters, named AMN1, AMN2, and AMN3 (ABCB for mycorrhization and nodulation), that are expressed early during infection by rhizobia and arbuscular mycorrhizal fungi. These ABCB transporters are strongly expressed in symbiotically infected tissues, including in root-hair cells with rhizobial infection threads and arbusculated cells. During nodulation, the expression of these genes is highly induced by rhizobia and purified Nod factors and is dependent on DMI3 but is not dependent on other known major regulators of infection, such as NIN, NSP1, or NSP2. During mycorrhization their expression is dependent on DMI3 and RAM1 but not on NSP1 and NSP2. Therefore, they may be commonly regulated through a distinct branch of the common symbiotic pathway. Mutants with exonic Tnt1-transposon insertions were isolated for all three genes. None of the single or double mutants showed any differences in colonization by either rhizobia or mycorrhizal fungi, but the triple amn1 amn2 amn3 mutant showed an increase in nodule number. Further studies are needed to identify potential substrates of these transporters and understand their roles in these beneficial symbioses.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Managing pain in HIV/AIDS: a therapeutic relationship is as effective as an exercise and education intervention for rural amaXhosa women in South Africa.

BACKGROUND: Pain is one of the most prevalent symptoms in people living with HIV/AIDS and is largely undermanaged. Both a peer-led exercise and education Positive Living programme (PL programme) and the PL programme workbook alone were previously found to be effective in reducing pain in urban amaXhosa Women Living With HIV/AIDS (WLWHA). A therapeutic relationship was hypothesised to have contributed to the efficacy of both interventions. The aim of the study was to determine the effectiveness of the PL programme and a therapeutic relationship, compared to a therapeutic relationship alone in managing pain amongst rural amaXhosa WLWHA on pain severity and pain interference, and secondary outcomes, symptoms of depression, health-related quality of life (HRQoL) and self-efficacy. METHODS: In this two-group, single-blind, pragmatic clinical trial with stratified convenience sampling, the PL programme and therapeutic relationship, was compared to a therapeutic relationship alone in rural amaXhosa WLWHA. The PL programme was a 6-week, peer-led intervention comprising education on living well with HIV, exercise and goal setting. The therapeutic relationship comprised follow-up appointments with a caring research assistant. Outcome measures included pain severity and interference (Brief Pain Inventory), depressive symptoms (Beck Depression Inventory), HRQoL (EuroQol 5-Dimensional outcome questionnaire) and self-efficacy (Self-efficacy for Managing Chronic Disease 6-Item Scale). Follow-up was conducted at 4, 8, 12, 24, and 48 weeks. Mixed model regression was used to test the effects of group, time, and group and time interactions of the interventions on outcome measures. RESULTS: Forty-nine rural amaXhosa WLWHA participated in the study: PL group n = 26; TR group n = 23. Both intervention groups were similarly effective in significantly reducing pain severity and interference and depressive symptoms, and increasing self-efficacy and HRQoL over the 48 weeks. A clinically important reduction in pain severity of 3.31 points occurred for the sample over the 48 weeks of the study. All of these clinical improvements were obtained despite low and suboptimal attendance for both interventions. CONCLUSIONS: Providing a therapeutic relationship alone is sufficient for effective pain management amongst rural amaXhosa WLWHA. These findings support greater emphasis on demonstrating care and developing skills to enhance the therapeutic relationship in healthcare professionals working with rural amaXhosa WLWHA. TRIAL REGISTRATION: PACTR; PACTR201410000902600, 30th October 2014; https://pactr.samrc.ac.za .

NIN Acts as a Network Hub Controlling a Growth Module Required for Rhizobial Infection.

The symbiotic infection of root cells by nitrogen-fixing rhizobia during nodulation requires the transcription factor Nodule Inception (NIN). Our root hair transcriptomic study extends NIN's regulon to include Rhizobium Polar Growth and genes involved in cell wall modification, gibberellin biosynthesis, and a comprehensive group of nutrient (N, P, and S) uptake and assimilation genes, suggesting that NIN's recruitment to nodulation was based on its role as a growth module, a role shared with other NIN-Like Proteins. The expression of jasmonic acid genes in nin suggests the involvement of NIN in the resolution of growth versus defense outcomes. We find that the regulation of the growth module component Nodulation Pectate Lyase by NIN, and its function in rhizobial infection, are conserved in hologalegina legumes, highlighting its recruitment as a major event in the evolution of nodulation. We find that Nodulation Pectate Lyase is secreted to the infection chamber and the lumen of the infection thread. Gene network analysis using the transcription factor mutants for ERF Required for Nodulation1 and Nuclear Factor-Y Subunit A1 confirms hierarchical control of NIN over Nuclear Factor-Y Subunit A1 and shows that ERF Required for Nodulation1 acts independently to control infection. We conclude that while NIN shares functions with other NIN-Like Proteins, the conscription of key infection genes to NIN's control has made it a central regulatory hub for rhizobial infection.

Barriers to implementing clinical trials on nonpharmacological treatments in developing countries: lessons learnt from addressing pain in HIV.

Pain affects over half of the people living with HIV/AIDS (LWHA), and pharmacological treatment has limited efficacy. Preliminary evidence supports nonpharmacological interventions. We previously piloted a multimodal intervention in amaXhosa women LWHA and chronic pain in South Africa with improvements seen in all outcomes, in both intervention and control groups. A multicentre, single-blind randomised controlled trial with 160 participants recruited was conducted to determine whether the multimodal peer-led intervention reduced pain in different populations of both male and female South Africans LWHA. Participants were followed up at weeks 4, 8, 12, 24, and 48 to evaluate effects on the primary outcome of pain, and on depression, self-efficacy, and health-related quality of life. We were unable to assess the efficacy of the intervention due to a 58% loss to follow-up (LTFU). Secondary analysis of the LTFU found that sociocultural factors were not predictive of LTFU. Depression, however, did associate with LTFU, with greater severity of depressive symptoms predicting LTFU at week 8 (P = 0.01). We were unable to evaluate the effectiveness of the intervention due to the high LTFU and the risk of retention bias. The different sociocultural context in South Africa may warrant a different approach to interventions for pain in HIV compared with resource-rich countries, including a concurrent strategy to address barriers to health care service delivery. We suggest that assessment of pain and depression need to occur simultaneously in those with pain in HIV. We suggest investigation of the effect of social inclusion on pain and depression.

Evidence for tryptophan being a signal molecule that inhibits conidial anastomosis tube fusion during colony initiation in Neurospora crassa.

Mycelial interconnectedness achieved by hyphal fusion has been hypothesized to facilitate the distribution and sharing of nutrients between different parts of a mycelium, especially when nutrients are heterogeneously distributed in the environment. However, the link between environmental nutrient availability and hyphal fusion is little understood. Here, we report that amino acids and extracellular pH regulate conidial anastomosis tube (CAT) fusion during colony initiation in Neurospora crassa. Quantitative analyses revealed that low extracellular pH and certain amino acids, particularly tryptophan, inhibit CAT fusion. Conidial germination was also inhibited by tryptophan but this inhibition was mitigated by the presence of other amino acids. This provides evidence for tryptophan having a role as a signal molecule that regulates CAT fusion. Tryptophan acts intracellularly because two amino acid permease mutants (Δmtr and Δaap-20) exhibited resistance against tryptophan-mediated inhibition of CAT fusion. Tryptophan and low pH did not significantly affect vegetative hyphal fusion in mature colonies, indicating that the latter is regulated in a different manner to CAT fusion.

Emergency department abdominal x-rays have a poor diagnostic yield and their usefulness is questionable.

OBJECTIVE: To determine the prevalence and nature of new clinically significant abdominal x-ray (AXR) findings and the proportion of patients receiving additional imaging in the emergency department (ED). METHODS: This was a retrospective audit of consecutive adult patients, who presented to a tertiary referral ED (annual census 70,000) between September and December 2008. Data were extracted from radiologist reports and the medical records of patients with new significant AXR findings. The electronic radiology record was further interrogated to determine which patients received additional imaging and whether this correlated with the original AXR findings. RESULTS: Of 997 cases that met the inclusion criteria, 121 (12.1%, 95% CI 10.2 to 14.4) and 43 (4.3%, 95% CI 3.2 to 5.8) had new clinically significant and insignificant AXR findings, respectively. Among the significant findings, the predominant diagnoses were bowel obstruction (72.7%), renal calculi (14.0%) and sigmoid volvulus (5.0%). Patient signs and symptoms were poorly associated with significant AXR findings. In all, 334 (33.5%, 95% CI 30.6 to 36.5) cases had additional imaging after the AXR. However, more patients with clinically significant AXR findings went on to have additional imaging (difference in proportions 23.0%, 95% CI 13.2 to 32.9, p<0.001). CONCLUSIONS: The yield for clinically useful information from the AXR is low and this investigation may be overused. Positive findings are associated mostly with bowel obstruction. As the proportion of patients ordered additional imaging was considerable, the utility of the preliminary AXR is questionable, especially in cases where the diagnosis is clear. Guidelines for AXR imaging are recommended to assist clinicians with investigation ordering.