Evolutionary biology of Fusarium oxysporum f.sp. cubense

Fusarium oxysporum Schlecht. is a cosmopolitan species complex that consists of both pathogenic and non-pathogenic members. The pathogenic members are subdivided into formae speciales, based on virulence to specific host species. More than 150 formae speciales have been described, of which F. oxysporum f.sp. cubense (E.F.Smith) Snyder and Hansen (Foc), causal agent of Fusarium wilt of banana, is regarded as one of the economically most important and destructive. According to phenotypic and genotypic markers, Foc has been classified into three races and 24 vegetative compatibility groups, and can be divided into a number of clonal lineages that roughly correspond with VCG groupings. In this thesis, we investigated the evolutionary relationships among VCGs using multi-gene sequencing and MAT genotyping. A PCR-RFLP fingerprint discriminating the Foc lineages and a PCR primer that identified Foc ‘subtropical’ race 4 isolates, was developed. Nine microsatellite markers (SSRs) were applied to a global population of Foc in order to investigate diversity not always detectable using sequencing data. Phylogenetic analysis of isolates representing Foc, various other formae speciales of F. oxysporum and non-pathogenic F. oxysporum of the genes encoding the translation elongation factor-1á (TEF), the mitochondrial small subunit (MtSSU), ribosomal RNA (rRNA), the repeated region encoded on the mitochondrion (MtR) and the intergenic spacer (IGS) gene regions separated these isolates into four clades, two of which included Foc. Within these two clades, Foc separated into six lineages that broadly corresponded to VCGs, while the non-pathogenic isolates of F. oxysporum grouped together in only one of the two clades, with an unknown Foc VCG isolate. The mating type of all isolates was determined and crosses were attempted between isolates harbouring MAT-1 and MAT-2 genes, without success. Cultural, morphological and pathogenic variation among isolates of Foc was unable to identify lineages as species. The separation of Foc isolates into two clades suggested that the banana pathogen evolved during two unrelated events. Factors such as horizontal gene transfer, however, might also have played a part in the pathogen’s evolution, as was evident from the divergent placement of some VCGs and lineages within the phylogenetic trees constructed. The inclusion of other formae speciales of F. oxysporum and non-pathogenic F. oxysporum isolates illustrated the great diversity that exists within the F. oxysporum complex. The inclusion of the Foc isolate of an unknown VCG suggests that the genetic diversity of Foc might be far greater than what is known and what was revealed in this study. The opposite mating types found in the respective lineages indicate a sexual origin for the Fusarium wilt fungus that could account for its polyphyletic nature. Within South Africa, Foc ‘subtropical’ race 4 is regarded the most important constrain to banana production. Conventional control practices for Fusarium wilt of banana are ineffective, and disease management relies heavily on the use of clean planting material and the early detection and isolation of the pathogen, in order to restrict spread to unaffected areas. Identification of Foc typically involves vegetative compatibility assays and pathogenicity testing using a set of differential host cultivars. The development of a PCR-based method for the rapid and accurate identification of Foc ‘subtropical’ race 4 will, therefore, be of great importance. The lack of morphological variation between lineages of Foc, and between pathogenic and non-pathogenic members, as well as the unreliability in race identification in Foc, makes the use of molecular tools a viable alternative. Following DNA isolation, PCR and sequencing of the MtR, the DNA sequence data revealed an 8-bp insertion that was subsequently targeted for the design of a Foc ‘subtropical’ race 4-specific primer. Isolates were positively identified as Foc ‘subtropical’ race 4 with the amplification of an 800-pb fragment. The development of the Foc ‘subtropical’ race 4 primer will aid in rapid and accurate detection of the Fusarium wilt pathogen of banana. The population structure defined according to SSR data of a global population of 239 Foc isolates resembled the structure defined according to multi-gene phylogeny, with some exceptions. Measures of gene and genotypic diversity unequivocally supported the opinion that Asia is the centre of origin of Foc. The presence of unique genotypes in all geographically-defined Foc populations could potentially indicate their evolution outside the centre of origin, although this is highly unlikely. The absence of certain genotypes from the Asian population was either due to insufficient and selective sampling, or it demonstrated the effects of clonal selection in combination with adaptation to the forces of geographic isolation and environmental changes over time. The worldwide collection of Foc mostly consisted of six over represented genotypes, thereby providing support for a clonal genetic structure. It was, however, not possible to reject the hypothesis of a recombining population for the populations representing isolates of Lineage V. The implication of recombination within some Foc lineages may be due to unobserved sexual reproduction in nature or the historical association with a sexual ancestor. When one considers diversity within and among genotypes, a specific genotype was mostly associated with only one or two Foc VCGs, therefore indicating that vegetative compatibility determination, in combination with phylogenetic analyses, is a powerful tool for characterizing isolates causing Fusarium wilt of banana. Results from this study, in combination with the multi-gene phylogeny, clearly indicated the presence of unrelated lineages that most probably represent cryptic species. Copyright 2008, University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. Please cite as follows: Fourie, G 2008, Evolutionary biology of Fusarium oxysporum f.s.p. cubense, MSc dissertation, University of Pretoria, Pretoria, viewed yymmdd E1216/gmDissertation (MSc)--University of Pretoria, 2008.Microbiology and Plant Pathologyunrestricte

Pantoea bathycoeliae sp. nov and Sodalis sp. are core gut microbiome symbionts of the two-spotted stink bug

DATA AVAILABILITY STATEMENT : The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found at: https://www.ncbi.nlm.nih.gov/genbank/, BioProject PRJNA972719 and PRJNA987703.Stink bug species (Pentatomoidea superfamily) have developed an interdependence with obligate bacterial gut symbionts in specialized midgut crypts (M4 sub-region). Species of the Enterobacteriaceae family (predominantly Pantoea) are vertically transferred to their offspring and provide nutrients that cannot be obtained from plant sap food sources. However, the bacteria in the other gut compartments of stink bugs have rarely been investigated. The two-spotted stink bug, Bathycoelia distincta, is a serious pest of macadamias in South Africa. Nothing is currently known regarding its gut microbiome or how symbionts are transferred between insect generations. In this study, the consistency of B. distincta gut bacteria across geographic locations and life stages was determined with 16S rRNA metabarcoding, considering both the M4 and other gut compartments. A novel Pantoea species was found to be the primary M4 gut symbiont and is vertically transferred to the offspring. The other gut compartments had a low bacterial diversity and genera varied between stink bug populations but a Sodalis species was prominent in all populations. Sequence data of the M4 compartment were used to produce high-quality metagenome-assembled genomes (MAGs) for the Pantoea and Sodalis species. Functional analyses suggested a similar role in nutrient provision for the host, yet also unique metabolites produced by each species. The Sodalis sp. also had additional traits, such as secretion systems, that likely allowed it to establish itself in the host. The Pantoea species was described as Pantoea bathycoeliae sp. nov based on the rules of the SeqCode.The Forestry and Agricultural Biotechnology Institute at the University of Pretoria and the South African Macadamia Association (SAMAC).http://www.frontiersin.org/Microbiologyam2024BiochemistryGeneticsMicrobiology and Plant PathologySDG-15:Life on lan

Body composition and intake of nutrients associated with bone metabolism in young adolescents in a peri-urban setting

OBJECTIVE : The aim was to describe the anthropometry, bone mineral content (BMC), bone mineral density (BMD), dietary calcium intake and 25(OH)D3 levels in 11- and 12-year-old children in a peri-urban area. DESIGN : A cross-sectional, descriptive study in the quantitative domain was undertaken. SETTING : Bronkhorstspruit, Gauteng, South Africa. SUBJECTS : Children, conveniently selected, were assessed in two groups. The first group comprised 70 children. From the 70 children, 20 children were conveniently selected to form a sub-sample (n = 20). OUTCOME MEASURES : Anthropometric data (weight, height) and dietary data (three quantified multi-pass 24-hour recalls). Children in the sub-sample additionally underwent body composition assessment (dual-energy X-ray absorptiometry; DXA scan) and a finger prick for 25(OH)D3. RESULTS : BMI and body composition data (body fat mass and lean fat mass) showed that the girls exceeded the boys in all measurements. The girls had a non-significantly higher BMD and BMC than the boys. The mean and median values for 25(OH)D3 were lower than the reference range values. Dietary intake results showed that the children had a sufficient macronutrient intake, but a deficient intake of calcium, phosphate and vitamin D. The sub-sample had a mean vitamin D intake of 3.2 mcg. CONCLUSION : The girls exceeded the boys in all the anthropometric and body composition measurements. The calcium and vitamin D intake of the children were of concern. There were no significant differences or relationships in the bone measurements and vitamin D status between the boys and girls.https://medpharm.tandfonline.com/loi/ojcn20am2019Human Nutritio

First report of Botrytis cinerea causing flower blight on macadamia in South Africa

DATA AVAILABILITY STATEMENT : The data that support the findings of this study are available from the corresponding author upon reasonable request.Macadamia cultivation plays significant role in the economy of South Africa. Despite its importance, the industry grapples with disease-related challenges, notably flower blight, which threatens substantial economic losses by affecting yield and nut quality. In 2022, diagnostic services at the Agricultural Research Council and the Forestry and Agricultural Biotechnology Institute received macadamia flowers displaying blight symptoms. Employing two methods for fungal isolation, 25 isolates were obtained which were classified into one group based on morphological characteristics. DNA analysis identified the isolates as Botrytis cinerea. The pathogenicity testing was conducted on macadamia flowers to confirm Koch's postulates. This is the first report of B. cinerea affecting Macadamia integrifolia Maiden & Betche in South Africa, underlining its potential threat to the industry.South African Macadamia industry (SAMAC).http://www.wileyonlinelibrary.com/journal/jphhj2024BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologySDG-01:No povertySDG-02:Zero Hunge

Biology and pathogenicity of fungi causing husk rot of macadamia in South Africa

Fungal diseases of macadamia fruit in South Africa have the potential to cause notable economic damage in this rapidly growing industry. To improve our understanding of the species involved in husk rot in macadamia orchards, a survey was conducted over two consecutive growing seasons to identify Colletotrichum, Diaporthe and Calonectria spp. that are associated with husk rot-infected macadamia fruits, and to investigate the occurrence of these fungi in asymptomatic and symptomatic fruits at the four stages of fruit development. Of the 425 fungal isolates obtained from the survey, Colletotrichum and Diaporthe were the most frequently isolated genera confirming the important role that these causal agents play in the husk rot epidemics. The detection of Calonectria species was low, only from symptomatic fruits and limited to a few locations in the main macadamia-producing provinces in South Africa. Colletotrichum and Diaporthe species were detected throughout the season at different stages of fruit development and in both symptomatic and asymptomatic fruits. The study confirmed that three fungal pathogens cause husk rot of macadamia in South Africa, with Colletotrichum and Diaporthe species that may have a latent phase in macadamia fruit. Studies of the growth characteristics of the husk rot pathogens revealed varied optimal growth temperatures, which may influence their prevalence in the different provinces in South Africa where macadamia is grown. The significance of the varied prevalence and biology of the causal agents in husk rot epidemics are discussed, which may be helpful management strategies.Macadamias South Africa NPC (SAMAC), the University of Pretoria, the Forestry and Agricultural Biotechnology Institute (FABI) and the DSI-NRF Centre of Excellence in Plant Health Biotechnology. Open access funding provided by University of Pretoria.https://link.springer.com/journal/10658hj2024Biochemistry, Genetics and Microbiology (BGM)Forestry and Agricultural Biotechnology Institute (FABI)SDG-02:Zero HungerSDG-15:Life on lan

Life history traits of the Pentatomidae (Hemiptera) for the development of pest management tools

Knowledge of the biology of a pest is essential for building sustainable management programmes. Pentatomidae have a hemimetabolous life cycle with egg, nymphal, and adult life stages, which differ in morphological, ecological, and behavioural traits. Some of these traits, such as mating behaviour, pheromones (alarm and aggregation pheromones) and the acquisition of gut symbionts can be targeted for pest management strategies. Here, we review the available literature on these life history traits of the Pentatomidae with potential for use in management programmes. Pheromone-mediated aggregation and the disruption of symbiont acquisition are two important targets for Pentatomidae control. Other traits such as the use of alarm pheromones for enhancing natural enemies and substrate-borne vibration for mating disruption deserve further consideration. Colour vision and flight ability are still poorly studied, despite their potential importance for stink bug management.The University of Pretoria, the Forestry and Agricultural Biotechnology Institute (FABI), the DSI-NRF Centre of Excellence in Plant Health Biotechnology, Macadamia South Africa NPC (SAMAC) and NRF Thuthuka.https://www.mdpi.com/journal/forestsam2024BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologyZoology and EntomologySDG-15:Life on lan

Lethal and sublethal effects of insecticides on Bathycoelia distincta (heteroptera: Pentatomidae)

DATA AVAILABILITY STATEMENT: The data that support the findings of this study are available from the corresponding author upon reasonable request.Bathycoelia distincta is one of the most dominant stink bug pests associated with macadamia orchards in South Africa. Understanding the toxicity and sublethal effects of insecticides on this pest is essential for its effective management. This study tested four commercial insecticide formulations, consisting of one organophosphate (acephate) and three pyrethroids (lambda-cyhalothrin, beta-cyfluthrin and tau-fluvalinate). The toxicity of these insecticides and their behavioural effects on mobility were assessed. The sublethal effects of lambda-cyhalothrin on the biological parameters of parent B. distincta (F0) and offspring generations (F1) were also determined by treating B. distincta adults with sublethal concentrations (LC10 and LC30). In toxicity bioassays, acephate was more toxic to B. distincta than lambda-cyhalothrin, beta-cyfluthrin and tau-fluvalinate. Behavioural changes were only observed in bugs exposed to pyrethroids, resulting in an increase in the distance walked and decrease of angular velocity. In the F0 generation, LC30 reduced the fecundity whereas the LC10 and LC30 accelerated development of the F1 generation. These results suggest that pyrethroids may enhance the dispersal of this pest and stimulate the growth of offspring populations. Further experiments should be conducted to confirm these results and understand the mechanism of action of pyrethroids on B. distincta.https://www.africanentomology.com/BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologySDG-02:Zero HungerSDG-15:Life on lan

Genetic diversity of the two-spotted stink bug Bathycoelia distincta (Pentatomidae) associated with macadamia orchards in South Africa

The South African macadamia industry is severely affected by a complex of stink bugs, dominated by the two-spotted stink bug, Bathycoelia distincta Distant (Pentatomidae). This species was first discovered during the spring of 1984 in the Limpopo province. Although considerable effort has been spent trying to manage this pest, it continues to be a pest of concern for the macadamia industry. Information on the genetic diversity of this species is lacking, despite the potential relevance of such information for management strategies. The present study aimed to characterise the genetic diversity of B. distincta populations in South Africa. The Cytochrome c Oxidase Subunit 1 (COI) and cytochrome b (Cytb) gene regions were sequenced from individuals collected from the three main regions of macadamia production over three different seasons (2018–2020). An overall high haplotype diversity (COI = 0.744, Cytb = 0.549 and COI+Cytb = 0.875) was observed. Pairwise mean genetic distance between populations from each region varied from 0.2–0.4% in both datasets, which suggests the absence of cryptic species. The median joining network for both datasets consisted of one or two central haplotypes shared between the regions in addition to unique haplotypes observed in each region. Finally, low genetic differentiation (FST < 0.1), high gene flow (Nm > 1) and the absence of a correlation between genetic and geographic distance were estimated among populations. Overall, these results suggest that the B. distincta populations are not structured among the areas of macadamia production in South Africa. This might be due to its ability to feed and reproduce on various plants and its high dispersal (airborne) between the different growing regions of the country along with the rapid expansion of macadamia plantations in South Africa.SUPPORTING INFORMATION : FIGURE S1. Isolation by distance of Bathycoelia distincta populations for COI marker (Mantel test, r = 0.731, P > 0.05). FIGURE S2. Isolation by distance of Bathycoelia distincta populations for Cytb marker (Mantel test, r = 0.310, P > 0.05). FIGURE S3. Isolation by distance of Bathycoelia distincta populations for COI+Cytb combined marker (Mantel test, r = 0.669, P > 0.05). TABLE S1. List of the individual for each haplotype generated in the study for the COI marker. TABLE S2. List of the individual for each haplotype generated in the study for the Cytb marker. TABLE S3. List of the individual for each haplotype generated in the study for the COI +Cytb combined markers.The University of Pretoria, the Forestry and Agricultural Biotechnology Institute (FABI), the Centre for Excellence in Plant Biotechnology (CPHB), Macadamia South Africa NPC (SAMAC) and NRF Thuthuka.http://www.plosone.orgdm2022BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologyZoology and Entomolog

Re-use of seedling containers and Fusarium circinatum association with asymptomatic Pinus patula planting stock

Fusarium circinatum is a pathogen causing serious post-planting mortality of Pinus patula seedlings in southern Africa. Containerised planting stock that is asymptomatic but associated with F. circinatum in the nursery is thought to be the cause of this problem. The aim of this study was to determine if re-use of seedling containers could be a source of inoculum resulting in asymptomatic planting stock and increased post-planting mortality of P. patula. Two experiments were conducted in successive years comparing nursery cull of symptomatic seedlings, seedling growth, association of F. circinatum with asymptomatic seedlings and post-planting mortality for crops raised in re-used containers, with and without sanitation, and factory-new containers. Each experiment consisted of a nursery production trial followed by out-planting into pots to assess post-planting mortality. Our results show that re-use of containers without sanitation increases the cull of symptomatic seedlings, incidence of F. circinatum associated with asymptomatic seedlings and post-planting mortality compared with the re-use of containers after steam sanitation or factory-new containers. Growth of asymptomatic seedlings was unaffected by container treatment or association with F. circinatum and in the absence of wilt symptoms the root system did not exhibit typical discolouration. Watering frequency did not influence post-planting mortality in pots. The comparison of two open-pollinated seed mixes of P. patula that, based on seedling stem inoculation screening, represented susceptible and tolerant material did not show differences in nursery cull or post-planting mortality. This work demonstrated that natural contamination of re-used containers can be a primary source of inoculum producing asymptomatic seedlings associated with F. circinatum that will succumb to the pathogen after field planting. The process of seedling infection, apparent latent infection in the seedling and expression of disease after planting needs greater understanding to improve nursery hygiene measures to control this disease.Forestry South Africa.http://www.tandfonline.com/loi/tsfs20hb201

Genetic diversity and population structure analysis reveals the unique genetic composition of South African selected macadamia accessions

Macadamia nuts are known globally for their high quality and economic value. Global macadamia commercial nut production amounts to 60,000 metric tonnes and is increasing steadily. South Africa is the leading producer with 29% of worldwide kernel production. Commercial macadamia germplasm was originally selected from a small genepool (mainly Macadamia integrifolia species) from a limited geographic distribution in Australia. These accessions were subsequently bred, cloned and exported across the world to start local macadamia industries. The South African macadamia industry was established with pre-commercial and commercial macadamia from different parts of the world, and local selections were also performed. Many of these accessions have unique genetic compositions that have not been characterized yet. We used 13 nuclear microsatellite markers to study the genetic diversity and structure of macadamia germplasm cultivated in South Africa. We compared four groups of accessions including 31 originating from the Hawaiian Agricultural Experimental Station (HAES), 19 from Australia (AUS), two from California and one from Israel (OTH), 31 from South Africa’s locally selected accessions (SA) and 26 from two local Farmers (FARM). We used STRUCTURE, PCoA and neighbour-joining phylogenetic analyses to show that the South African selected accessions include diverse hybrid genotypes with strong Macadamia tetraphylla composition, unlike the Hawaiian commercially released and Australian representative collections that mostly have M. integrifolia or hybrid composition. Our results suggest that the South African selections represent a unique and diverse set of germplasm for future macadamia improvement efforts that will benefit from genomic breeding technologies.The National Research Foundation (NRF) of South Africa and Macadamias South Africa.https://link.springer.com/journal/11295hj2023BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog