This report details a field research study identifying viral diseases affecting barley crops in South Korea. RT-PCR tests showed that samples were infected with both Barley yellow dwarf virus-PAS and Hordeum vulgare alphaendornavirus. The high occurrence of viruses during the spring season was likely due to suitable climate conditions for the distribution of viral vectors like aphids. A new endornavirus, named Hordeum vulgare alphaendornavirus Buan, was also identified that was 98% identical to a previously reported endornavirus in Korea. The study helps better understand the impact and prevalence of these viruses in barley crops in the Buan region.

1. UST Field Research Result Report
Approval
Person in Charge Team Head Dean President
Campus/ Major KRIBB/ biotechnology
Course
Program
Field Research (Internal)
Student Number 02013035
Semester
registered
3rd
Semester
Classification Advisor's Name Mesele Tilahun Belete
Subject Name Field Research A (Field trip of plant pathology)
Period/Time February – June 2021 Place
Plant engineering research
center (KRIBB) building no.
4209
Summarized Content
Identification of Viral Diseases Affecting Barley Crops in Spring Season, Buan Region of South
Korea
Abstract: The RT-PCR detecting result showed that barley yellow dwarf virus-PAS and Hordeum
vulgare alphaendornavirus virus was observed on the same barley sample. The occurrences of virus
during spring season was high even though it needs repeated assessment in the same season. This
might be happened by aphid distribution in region, due to current climate situations, the 2021spring
period could be a somewhat humid season, which probably increased distribution of vectors and, as a
result, virus occurrence and the same barley infected more than one virus. The partial nucleotide
amplified preexisting infecting Endornavirdae type, temporarily we named Hordeum vulgare
Endornaviruses Buan (HvEVB), which identified in Buan region, South Korea, has been detected by
RT-PCR. The viral genomic RNA amplified is 725bp and the Sequence comparison showed that
HvEVB 98% resembled with Hordeum vulgare endornavirus which reported in Korea 2016. This
finding indicated that HvEVB is a descendant of a genera of pre-existing endornavirus.
Key words: RT-PCR, Hordeum vulgare alphaendornavirus, Barley
INTRODCUTION: Barley (Hordeum vulgare L.) is a kind of cereal grass belonging to the family
Poaceae [1] are primarily mainly used for animal feed, human food, and malting. It widely
cultivated in West Asia and North Africa (WANA), where they are great sources of protein and
calories for a majority of the population [2] [3]. Based on demand, barley is the 4th
leading import
crop next to maize, rice, and wheat. Barley is commonly grown and eaten as processed and mixed
with rice in Korea , but it can also important to make noodles, bread, pasta, beverages, tea,
and oils [1]. Bacteria, fungi, and viruses are some of the pathogens that may attack barley. Mostly
Viruses poses a serious threat to the yields of barley both in terms of quality and quantity. Virus-
borne diseases cause great economic losses all around world, including crop failure, yield and quality
losses, and raised resource expenses related with disease prevention and control[4]. Several viruses
have been detected in cereal pants all around world, even though only a few are reported Barley
yellow dwarf virus PAV [5] , Barley stripe mosaic virus , Barley yellow mosaic virus, Barley mild
mosaic virus, Hordeum mosaic virus, Hordeum vulgare endornavirus [1], [6], [3].

2. In Korea, there haven't been many reports of viral infections in barley crops., and rare data on their
prevalence is available. The study's main goals were to figure out and determine what type of viruses
affect monocot crops such as barley in major growing areas in Korea, as well as their occurrence and
relevance.
Fig.2. Image of barley plants showing viral disease
symptom
Materials and Methods: Sample collection and field tour took place between April 14- 17, 2021,
when the plants have been before their reproductive phase. We were collect the sample having the
virus-like symptoms of leaf malformation and stunting. The collected sample in the region plants
exhibiting viral and viron symptoms have been chosen randomly at farming field. A total of 3 barley
farmers' fields were picked for sampling based on crop condition, growth stage and viral disease
symptoms estimation near a location Pungwol-r1, Buan, Republic of Korea. The samples were put
in explicitly labeled plastic bags and brought to Korean Research Institute of Bioscience and
Biotechnology(KRIBB) laboratory in building 6th
, 4th floor. For further studying, the collected
samples were kept in fine powder at -80 °C.
To further confirm the sample was weather infected with virus or not, 22 specific primer pairs from
the known sequence of complete genome 18 barley infected virus genera were designed using
primer3plus software based on its conserved sequence coat protein (CP) region, which were obtained
from comparing the available sequences of related genera through the corresponding isolates in
GenBank.
The Barley sample RNA was extracted from -80 °C stored fine powdered barley sample using a
WizPrep™ Plant RNA Kit (Wizbiosolutions, Republic of Korea) and cDNA were synthesized with
the 25-mer degenerate primer N25 using Revert Aid Reverse Transcriptase (Thermo Scientific,
Waltham, MA, USA), following to the manufacturer’s instructions. The cDNAs were amplified by
PCR with AccuPower ProFi Taq PCR PreMix kit (Bioneer, Republic of Korea) using that of 22 oligo
pair of primer according to the thermal profile: 95 °C for 5 min followed by 36 cycles at 95°C for 20
seconds, annealing in temperature appropriate to primers for 30 seconds and extension step at 68°C
for 1 minutes per 1 kb. The RT-PCR amplified products were analyzed by 1% gel electrophoresis
having EtBr staining and visualized under UV light. The PCR product have been purified by
QIAquick® PCR Purification Kit and then sequenced by Macro gene (Seoul, Korea).
Discussion and Result: The virus occurrences during spring season was relatively high. This might
be, by aphid distribution in several fields site, due to the current climate situations, the 2021spring
period was a somewhat humid, which probably increased vector distribution, as a result, virus spread
and the same barley infected by more than one virus. Barley yellow dwarf virus-PAS are transmitted
by aphids [2], and because virus spread is probably to occur when climate situation are suitable to
viral vector emergence and migration.
Using the extracted RNA from sample that brought in Buan, South Korea showed viral infections
similar to that of Hordeum vulgare alphaendornavirus (HvEV) and Barley yellow dwarf virus PAS

3. (BYDV-PAS) with expected sizes 725bp (Gene bank acc no. NC_028949.1 ) and 405bp (Gene bank
acc no. LC592174.1) respectively were amplified. For further confirmation HvEV PCR result was
purified by QIAquick® PCR Purification Kit & then sequencing performed by Macro gene (Seoul,
Korea), However, the second viral amplified result BYDV-PAS was not checked by sequencing.
As expected, annotation of HvEV analyses showed that 98.83% identical to endornaviral sequence of
Hordeum vulgare alphaendornavirus (Gene bank acc. No. MF979141.1), which reported in Apr-2016
in Korea. Therefore, we temporarily named it Hordeum vulgare alphaendornavirus Buan (HvEVB).
This HvEVB is plant-infecting viruses that is the member of genus Endornavirus in the family of
Endornaviridae and poses 14-17kb linear double-stranded RNA (dsRNA) genome and a persistent
lifestyle [6].
Conclusion: In conclusion, this study will help investigation and estimates of what viruses occurred
and infect barley plant in spring season and a more understanding of the effect of BYDV-PAS and
HvEV in Buan region, republic of Korea.
Reference
[1] Y. Jo, J. Y. Bae, S. M. Kim, H. Choi, B. C. Lee, and W. K. Cho, “Barley RNA viromes in six
different geographical regions in Korea,” Sci. Rep., vol. 8, no. 1, pp. 1–13, 2018, doi:
10.1038/s41598-018-31671-4.
[2] S. G. Kumari et al., “Identification of viral diseases affecting barley and bread wheat crops in
Yemen,” Australas. Plant Pathol., vol. 35, no. 5, pp. 563–568, 2006, doi: 10.1071/AP06061.
[3] F. Zhao et al., “The complete genomic sequence of a tentative new polerovirus identified in
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10.1007/s00705-016-2881-0.
[4] N. Nancarrow, M. Aftab, G. Hollaway, B. Rodoni, and P. Trębicki, “Yield losses caused by
barley yellow dwarf virus‐pav infection in wheat and barley: A three‐year field study in
south‐eastern Australia,” Microorganisms, vol. 9, no. 3, pp. 1–15, 2021, doi:
10.3390/microorganisms9030645.
[5] P. P. Ueng et al., “Nucleotide sequence analysis of the genomes of the MAV-PS1 and P-PAV
isolates of barley yellow dwarf virus characterized by restriction enzyme digestion and by
( ORFs ) which are similar to those identified from a BYDV isolate from Australia ( Vic-
PAV ). ,” pp. 487–492, 1992.
[6] T. Candresse et al., “Complete genomic sequence of barley (Hordeum vulgare) endornavirus
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