Australasian Plant Disease Notes
(2023) 18:16
https://doi.org/10.1007/s13314-023-00501-6
Detection and molecular characterization of a new bipartite
begomovirus infecting asystasia (Asystasia gangetica) in Benin
Republic in West Africa
Bahoya JA Lauryn1 · Ekwa Y Monono1 · Jean CM Piau1 · Boniface D Kashina2 · Vincent N Fondong3 · Walter N Leke1,3
Received: 5 March 2023 / Accepted: 27 April 2023
© The Author(s) under exclusive licence to Australasian Plant Pathology Society Inc. 2023
Abstract
Asystasia gangetica (A. gangetica) leaves exhibiting yellow mosaic symptoms were tested for the presence of begomovirus
infection. The putative full-length of a bipartite begomovirus components A and B molecules, respectively were amplified,
sequenced, and analyzed. This represents a distinct begomovirus species infecting Asystasia in Benin Republic in West Africa.
Keywords Acanthaceae: Asystasia · Geminiviridae: begomovirus · West Africa Asystasia virus
Asystasia gangetica (L.) T. Anderson, belongs to the family Acanthaceae which is an annual or perennial invasive
semi-woody dicot herb (weed) wildly distributed across
West and Central Africa and is used for food and medicine
(Irvine 1938).Yellow mosaic disease symptoms are commonly observed on Asystasia gangetica throughout West
and Central Africa (Fig. 1); these plants are also observed
to be heavily infested by whiteflies Bemisia tabaci (Genn.),
which are vectors of viruses in the genus Begomovirus (family Geminiviridae). This suggests that these yellow mosaic
symptoms may be caused by begomoviruses. Thus, in 2014,
we collected leaf samples randomly from three symptomatic
and three asymptomatic A. gangetica plants from Cotonou
(6o2213.05N and 2o2328.45E) in Benin Republic in West
Africa. The leaves were press-dried and stored until DNA
extraction. Total DNA was isolated as described by Shepherd
et al. (2008). The viral circular, ssDNA was enriched in each
of the samples by rolling cycle amplification (RCA) (GE
Healthcare Bio-Science, Piscataway, NJ). The RCA products
* Walter N Leke
walter.leke@fulbrightmail.org
1
Present Address: Institute of Agricultural Research
for Development (IRAD), Ekona, PMB 25, Buea,
Southwest Region, Cameroon
2
Department of Crop Protection, Ahmadu Bello University,
Zaria, Nigeria
3
Department of Biological Sciences, Delaware
State University, 1200 N. Dupont Highway, Dover,
DE 19901-2277, USA
were pooled for both the symptomatic plants (BNAsy58) and
asymptomatic plants (BN_C), respectively. The two samples
were subjected to Illumina sequencing (MisEquation 2 ×
150-bp configuration). The sequences were assembled using
SeqMan NGen software (DNASTAR, Madison, WI). The
putative full-length begomovirus sequences for BNAsy58
were 2,728-bp for DNA-A and 2, 661-bp for DNA-B. As
expected, no viral sequences were detected in BN_C from
asymptomatic plants.
The BLASTn (Altschul et al. 1990) and pairwise SDT
(Muhire et al. 2013) analyses of the full-length begomovirus sequences revealed the closest identity of BNAsy58
putative DNA-A (KT444609) (89.4%) and DNA-B
(KT444610) (93.8%) to the West African Asystasia virus1
(WAAV1) DNA-A and DNA-B, respectively (Wyant et al.
2015). Sequence analysis showed that BNAsy58 sequences
exhibit typical bipartite begomovirus genome organization. Thus, DNA-A contained four open reading frames
(ORFs): AV1 and AV2 in the virion sense strand, and AC1,
AC2, AC3, and AC4 in the complimentary sense strand.
Correspondingly, DNA-B contained two ORFs: BV1 in
the virion sense strand and BC1 in the complimentary
sense strand. No begomovirus-associated satellites (alphasatellite or betasatellite) genome and other virus-specific
contigs were present in the Illumina reads. Based on the
Illumina-derived viral sequences, the endonucleases SacI
(DNA-A) and ApaI (DNA-B) were used to digest the RCA
products of the three symptomatic and three asymptomatic
samples. This digestion yielded ~2.8-bp fragments for all
three samples from the symptomatic plants, in contrast no
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Australasian Plant Dis. Notes
(2023) 18:16
Fig. 1 Asystasia gangetica with
yellow mosaic symptoms, typical of begomovirus infection
such fragment was observed in samples collected from the
asymptomatic plants. The fragments were cloned into a similarly digested pGEM-5Zf (+) plasmid vector (Promega) and
sequenced bidirectionally by primer walking. The Sangerand Illumina derived asystasia begomovirus sequences were
100% identical for both DNA-A and DNA-B components.
Recombination Detection Program (RDP5) (Martin et al.
2020) analysis of the begomovirus sequences subjected to
phylogenetic analysis did not provide significant support for
recombination for the newly discovered West African Asystasia begomovirus species, based on a significant p-value of
<10-3 for a minimum of three RDP methods.
Fig. 2 Phylogenetic relationships between the DNA-A and B components of West Africa Asystasia virus3 (WAAV3) identified in the
Benin Republic in West Africa and related begomoviruses (abbreviations according to Brown et al. 2015). Trees constructed using
the neighbour-joining algorithm in MEGA 11. The begomovirus
sequences identified in this study are shown in bold. Horizontal lines
are proportional to the number of nucleotide substitutions per site.
Bootstrap values of ≥70% are shown at the major nodes. GenBank
Accession Nos. and isolate names for each virus are indicated. The
trees are rooted on yet another new bipartite begomovirus identified
infecting cotton in Benin Republic (Leke et al. 2016b)
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Australasian Plant Dis. Notes
(2023) 18:16
According to the recommendations of the International
Committee on Taxonomy of Viruses (Brown et al. 2015) and
the phylogenetic relatedness of the Benin asystasia begomovirus with the previously described asystasia begomoviruses
(Fig. 2), the results indicate that the begomovirus associated with asystasia yellow mosaic disease in Benin Republic
(Fig. 1), is a new bipartite begomovirus and has been named
by ICTV as West African Asystasia virus3 (WAAV3). West
African Asystasia virus1 and West African Asystasia virus2
have been reported to infect asystasia and cassava in West and
Central Africa (Wyant et al. 2015; Leke et al. 2016a; Bahoya
et al. in press). Surprisingly, West African Asystasia virus2,
reported to co-infect asystasia with West African Asystasia
virus1 (Wyant et al. 2015) was not detected in this present
study. The identification of WAAV3 which represents a new
begomovirus species suggests asystasia is a major host to
begomoviral complexes in West and Central Africa, which are
predominantly bipartite in nature. Elsewhere in Madagascar in
southern Africa, another begomovirus was identified infecting
asystasia, with bipartite genome organization, named Asystasia mosaic Madagascar virus (AMMGV) (De Bruyn et al.
2015), just as WAAV1 and WAAV3 in this study. This study
is the first to identify a begomovirus infecting asystasia in
the Benin Republic in West Africa, representing a different
begomovirus species. This study, together with previous studies strongly suggests that, apart from WAAV2, asystasia is
predominantly infected by bipartite begomoviruses.
Funding This research was partly funded by National Science Foundation award number IOS-1212576. Walter Leke was supported by the
Fulbright Foundation.
Declarations
Conflict of interest The authors declare that they have no conflict of interest.
Page 3 of 3
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Brown JK, Zerbini FM, Navas-Castillo J, Moriones E, Ramos-Sobrinho
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