Using qPCR and microscopy to assess the impact of harvesting and weather conditions on the relationship between <i>Alternaria alternata</i> and <i>Alternaria</i> spp. spores in rural and urban atmospheres

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Abstract

Alternaria is a plant pathogen and human allergen. Alternaria alternata is one of the most abundant fungal spores in the air. The purpose of this study was to examine whether Alternaria spp . spore concentrations can be used to predict the abundance and spatio-temporal pattern of A. alternata spores in the air. This was investigated by testing the hypothesis that A. alternata dominates airborne Alternaria spp. spores and varies spatio-temporally. Secondarily, we aimed at investigating the relationship between airborne Alternaria spp. spores and the DNA profile of A. alternata spores between two proximate (~ 7 km apart) sites. These were examined by sampling Alternaria spp. spores using Burkard 7-day and cyclone samplers for the period 2016–2018 at Worcester and Lakeside campuses of the University of Worcester, UK. Daily Alternaria spp. spores from the Burkard traps were identified using optical microscopy whilst A. alternata from the cyclone samples was detected and quantified using quantitative polymerase chain reaction (qPCR). The results showed that either A. alternata or other Alternaria species spores dominate the airborne Alternaria spore concentrations, generally depending on weather conditions. Furthermore, although Alternaria spp. spore concentrations were similar for the two proximate sites, A. alternata spore concentrations significantly varied for those sites and it is highly likely that the airborne samples contained large amounts of small fragments of A. alternata. Overall, the study shows that there is a higher abundance of airborne Alternaria allergen than reported by aerobiological networks and the majority is likely to be from spore and hyphal fragments.

Supplementary Information

The online version contains supplementary material available at 10.1007/s00484-023-02480-w.

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The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments.

Stephen A. Bustin, Vladimir Benes, Jeremy A. Garson … (2009)

Currently, a lack of consensus exists on how best to perform and interpret quantitative real-time PCR (qPCR) experiments. The problem is exacerbated by a lack of sufficient experimental detail in many publications, which impedes a reader's ability to evaluate critically the quality of the results presented or to repeat the experiments. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency. MIQE is a set of guidelines that describe the minimum information necessary for evaluating qPCR experiments. Included is a checklist to accompany the initial submission of a manuscript to the publisher. By providing all relevant experimental conditions and assay characteristics, reviewers can assess the validity of the protocols used. Full disclosure of all reagents, sequences, and analysis methods is necessary to enable other investigators to reproduce results. MIQE details should be published either in abbreviated form or as an online supplement. Following these guidelines will encourage better experimental practice, allowing more reliable and unequivocal interpretation of qPCR results.

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Alternaria section Alternaria: Species, formae speciales or pathotypes?

J.H.C. Woudenberg, M.F. Seidl, J.Z. Groenewald … (2015)

The cosmopolitan fungal genus Alternaria consists of multiple saprophytic and pathogenic species. Based on phylogenetic and morphological studies, the genus is currently divided into 26 sections. Alternaria sect. Alternaria contains most of the small-spored Alternaria species with concatenated conidia, including important plant, human and postharvest pathogens. Species within sect. Alternaria have been mostly described based on morphology and / or host-specificity, yet molecular variation between them is minimal. To investigate whether the described morphospecies within sect. Alternaria are supported by molecular data, whole-genome sequencing of nine Alternaria morphospecies supplemented with transcriptome sequencing of 12 Alternaria morphospecies as well as multi-gene sequencing of 168 Alternaria isolates was performed. The assembled genomes ranged in size from 33.3–35.2 Mb within sect. Alternaria and from 32.0–39.1 Mb for all Alternaria genomes. The number of repetitive sequences differed significantly between the different Alternaria genomes; ranging from 1.4–16.5 %. The repeat content within sect. Alternaria was relatively low with only 1.4–2.7 % of repeats. Whole-genome alignments revealed 96.7–98.2 % genome identity between sect. Alternaria isolates, compared to 85.1–89.3 % genome identity for isolates from other sections to the A. alternata reference genome. Similarly, 1.4–2.8 % and 0.8–1.8 % single nucleotide polymorphisms (SNPs) were observed in genomic and transcriptomic sequences, respectively, between isolates from sect. Alternaria, while the percentage of SNPs found in isolates from different sections compared to the A. alternata reference genome was considerably higher; 8.0–10.3 % and 6.1–8.5 %. The topology of a phylogenetic tree based on the whole-genome and transcriptome reads was congruent with multi-gene phylogenies based on commonly used gene regions. Based on the genome and transcriptome data, a set of core proteins was extracted, and primers were designed on two gene regions with a relatively low degree of conservation within sect. Alternaria (96.8 and 97.3 % conservation). Their potential discriminatory power within sect. Alternaria was tested next to nine commonly used gene regions in sect. Alternaria, namely the SSU, LSU, ITS, gapdh, rpb2, tef1, Alt a 1, endoPG and OPA10-2 gene regions. The phylogenies from the two gene regions with a relatively low conservation, KOG1058 and KOG1077, could not distinguish the described morphospecies within sect. Alternaria more effectively than the phylogenies based on the commonly used gene regions for Alternaria. Based on genome and transcriptome comparisons and molecular phylogenies, Alternaria sect. Alternaria consists of only 11 phylogenetic species and one species complex. Thirty-five morphospecies, which cannot be distinguished based on the multi-gene phylogeny, are synonymised under A. alternata. By providing guidelines for the naming and identification of phylogenetic species in Alternaria sect. Alternaria, this manuscript provides a clear and stable species classification in this section.

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Pollen monitoring: minimum requirements and reproducibility of analysis

C. Saldaña-Galán, M M Smith, M Thibaudon … (2014)

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Author and article information

Contributors

Godfrey Philliam Apangu:

ORCID: http://orcid.org/0000-0002-0707-8754

godfrey.apangu@rothamsted.ac.uk

Journal

Journal ID (nlm-ta): Int J Biometeorol

Journal ID (iso-abbrev): Int J Biometeorol

Title: International Journal of Biometeorology

Publisher: Springer Berlin Heidelberg (Berlin/Heidelberg )

ISSN (Print): 0020-7128

ISSN (Electronic): 1432-1254

Publication date (Electronic): 16 May 2023

Publication date PMC-release: 16 May 2023

Publication date (Print): 2023

Volume: 67

Issue: 6

Pages: 1077-1093

Affiliations

[1 ]GRID grid.189530.6, ISNI 0000 0001 0679 8269, School of Science and the Environment, , University of Worcester, ; Henwick Grove, WR2 6AJ Worcester, UK

[2 ]GRID grid.418374.d, ISNI 0000 0001 2227 9389, Present Address: Protecting Crops and the Environment, , Rothamsted Research, ; West Common, Harpenden, Hertfordshire AL5 2JQ UK

[3 ]GRID grid.454322.6, ISNI 0000 0004 4910 9859, Present Address: Department of Urban Greening and Vegetation Ecology, , Norwegian Institute of Bioeconomy Research, ; Ås, Norway

[4 ]GRID grid.1038.a, ISNI 0000 0004 0389 4302, Present Address: Edith Cowan University, ; 270 Joondalup Drive, Joondalup, WA 6027 Australia

[5 ]GRID grid.7048.b, ISNI 0000 0001 1956 2722, Present Address: Department of Environmental Science, , Aarhus University, ; Frederiksborgvej 399, 4000 Roskilde, Denmark

Author information

Godfrey Philliam Apangu http://orcid.org/0000-0002-0707-8754

Article

Publisher ID: 2480

DOI: 10.1007/s00484-023-02480-w

PMC ID: 10267013

PubMed ID: 37191729

SO-VID: dc6d6dc2-e689-481d-941b-33bd64692e49

License:

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.