MINISTRY OF EDUCATION MINISTRY OF AGRICULTURE

AND TRAINING AND RURAL DEVELOPMENTVIET NAM ACADEMY OF AGRICULTURAL SCIENCES
LE MAI NHAT STUDY ON GREENING IN CITRUS FRUIT TREE IN SOME NORTHERN
PROVINCES OF VIET NAM AND RECOMMENDATIONS FOR
PREVENTIVE AND CONTROLLABLE MEASURES

Branch of study: Plant protection
Code: 62 62 01 12 SUMMARY OF AGRICUTURAL DOCTORAL THESIS



Critic 3: The thesis is defended at Council for Thesis Assessment at
Institutional level, held at:

Viet Nam Academy of Agricutural Sciences
At hours, day month year 2014
This thesis can be referred to at:
1. Viet Nam National Library
2. The Library Viet Nam Academy of Agricutural Sciences

1

U
1. Rationale of the research
Citrus (orange, mandarin, lime, pomelo…) is an important agricultural fruit crop
in Vietnam. In 2011, the total of citrus grown area in Vietnam has reached 124,057
ha, comprising orange and mandarin 70,300 ha, pomelo 45,000 ha, lime 18,000 ha.
Citrus produce is not only for domestic consumption but also is a high value export

could be used to develop an effective control system, partly contribution to the
development, conservation and sustainability of citrus industry in Vietnam.
4. Subject and scope of research
4.1. Research subject
- Bacteria Liberibacter asiaticus – a causative agents of greening in citrus and disease
transmission psyllid Diaphorina citri Kuwayama
2

- Selected local and import fruit crops which are grown in some provinces in
Northern Vietnam, crops in Rutaceae and are hosts of L. asiaticus that causes VLG in
citrus.
4.2. Scope of research
This research studies VLG transmissions, factors influencing to the
transmission and development of VLG within the ecological of Northern provinces of
Vietnam. The research identifies bacteria of L.asiaticus causing VLG in citrus and
their distributions as well as studies effective measures to prevent and control VLG in
some Northern provinces.
5. New contributions of dissertation
- Providing a scientific source on popularity, symptom, and host of VLG in citrus in
Northern Vietnam.
- Identifying bacteria causing VLG belonging type I and II in present citrus crops in
production areas of Northern Vietnam for the first time.
- Finalizing and perfecting shoot tip grafting to clean VLG and other viruses,
contributing to complete technical procedures for virus-free grafted planting
materials.
- Evaluating VLG-resistant capability of some grafted local and import rootstocks and
recommending some measures to prevent VLG re-infection.
6. Structure of thesis
This dissertation comprises 146 pages, including the introduction 4 pages; Chapter 1.
Scientific background and literature review (36 pages); Chapter 2. Research materials,

1.2. Review of international literature
1.2.1. VLG’s symptoms, history and distribution
VLG was firstly recorded from 1929 in China with different names such as
Huanglongbing or yellow shoot in South China. The disease is then documented in
1943, Greening in South Africa (1947), Likubin (Tawain, 1951), Leaf mottle yellows
in the Philippines, huanglongbing in Thailand (1960), Dieback in India, greening in
Myanmar (1970), Vein Phloem degeneration in Indonesia (1980), greening in Vit
Nam, Laos, Cambodia (1980), Huanglongbing in Iriomote, Okinawa (1994),
Okinawa Island, Japan (2003), in Tokunoshima, Kagoshima, Japan (2003),
Huanglongbing in Brazil (2004) and in Florida, the United States (2005). Presently,
the disease has not been found in Australia yet.
1.2.2. Diagnose and appraise VLG
Identifying the causative agent by electron microscope: The VLG’s causative
bacteria is observed by electron microscope. The VLG bacteria typically has a tinny
long stick shape and changes from young to mature cells (Huang, 1987).
Identifying the causative agent by bio-molecular method: This method uses specific
primers to diagnose VLG from fresh sample and insect vector (Hung et al., 1999,
Jagoueix et al., 1996; A. Hocquellet 1997; Subandiyah et al., 2000).
Researches on type of VLG’s bacteria: Su, (2008) had identified types of VLG’s
bacteria in Taiwan by a combined method which uses plant indicator and other
selected fruit trees with biotechnology. Since then, there are four types of VLG
bacteria have been identified. Different types cause different symptoms on those fruit
trees.
1.2.3. Researches on pathology and ecology of VLG
VLG spreads very fast and reaches more than 95% in a period from 3 to 9
years after the first symptoms found (Matsumoto et al., 1961; Aubert et al., 1988,
Bové et al., 2000b; and Gottwald, 2005).
VLG spreads out through propagation and vectored insects (Diaphorina citri
Kuwayama) in a sustainable way (Halbert and Manjunath, 2004; Pluke et al., 2008).
Skelley & Hoy (2004) have applied a method of multiplying psyllid on

1.3.3. Research on pathology
Hà Minh Trung et al, (1995) have conducted researches on VLG’s transmission
through grafting, grafted budwood and vector.
Ngô Vnh Vin et al., (2009), Nguyn Vn Hoà et al., (2012) have collected and
primarily assessed the resistant capability of selected trees of Rutaceae.
1.3.4. Research on disease management
Currently, technical guidelines and procedures for shoot tip grafting have been
improving to increase the shoot tip grafted tree’s ability of disease cleanliness and
survival. Net-house systems are established to maintain elite trees and clean disease
for shoot tip grafted materials. Disease management measures are simultaneously
implemented from breeding and selection stages to improve planting materials to
cultivational, biological and chemical techniques to prevent and control disease
vectors.
Chapter 2
RESEARCH MATERIAL, CONTENT AND METHODOLOGY
2.1. Research location and period
5

Research location
- Division of Plant Pathology, Plant Protection Research Institute, ông Ngc,
 Liêm, Hà Ni
- Laboratory in National Taiwan University of Science and Technology
- Nghi Diên, Nghi Lc, Ngh An and other Northern provinces of Vietnam
Research period: January, 2010 to October, 2013
2.2. Research materials
MS environment (Murashige and Skoog), BAP, plant growth regulators used in
shoot tip grafting. Seed for rootstocks (Three leaves orange, sour pomelo, chp,
Green Orange Citrus reticulata × maxima, Cleopatra mandarin). Chemicals used in
identifying VLG causative agents.
Free disease citrus trees (Cam Canh, Xã oài, Din pomelo, Eureka lime, Thanh

6

2.4.3. Methods for researching on identification of VLG’s host and VLG’s
bacteria distribution on citrus in some Northern provinces of Vietnam
Following methods of Su, (2008), Hung et al., 1999 and specific primers in
order to identify causative bacteria in citrus trees of Tomimura et al., 2010.
2.4.4. Methods for researching on some techniques to prevent and control of
VLG
Shoot tip grafting: Following methods of Su and Chu (1984). Procedures
including preparation of rootstock and shoot tip, first grafting, invitro caring. After
that, second grafting and transplanting trees out of nursery. In this study, several
milestones have been done and improved to enhance survival and disease cleanliness
rates by the technique in order to produce disease free materials.
Disease-free planting material techniques: To be conducted in a complete
procedure in three-level net-house, including: container’s substrate and seed sowing;
Soil-less planting material production techniques; Technical and budding method.
Method for assessment of resistance of rootstock which being used in VLG
planting material production: Following methods of Kranz (1988) and Bowen (2004)
Method for studying of anti-reinfection of VLG in the field: Following common
methods on plant protection in citrus trees.
2.5. Method for processing of experiment data: Experimental data are processed
by statistical processor MS.Exel, 2007 and IRRISTAT VERSION 5.0
Chapter 3
RESEARCH RESULTS AND DISCUSSIONS
3.1. VLG’s popularity and symptoms
3.1.1. Current use of citrus varieties in Hà Ni and Hòa Bình
In last years, citrus trees have been grown rapidly, however, growers have not
known yet about the source of disease latent in the trees. Result on the survey on
original and quality of planting material shows that, in Hanoi, 46.67% of planting
materials are self-produced, 40.0% are unidentified of original, only 13.33% are

samples infected.
3.1.4. Degree of VLG infection in integration with other virus, viroid diseases in
citrus crops
Other virus diseases have promoted a faster and more severe process of
degradation in citrus orchards. VLG and tristeza have severely damaged all citrus
crops in the survey. The rate of VLG infection is from 60.0 to 86.0% of the total
samples surveyed; tristeza is from 27.91 to 37.78%. Especially, in our surveys, a
disease called exocortis has been identified on orange for the first time with a rate of
detection of 14.28% in the total samples collected and a disease named tatter leaf on
mandarin with an infected rate of 9.68% in the total samples collected (table 3.5).
Table 3.5. Result of identification of presence of VLG and some other virus
diseases in citrus crops on the same sample by molecular
(at Plant Protection Research Institute, 2010 – 2012)
Citrus
crops
Scientific name Total
samples
Rate (%) sample infected
VLG

CTV

CTLV

CEVd

Orange Citrus sinensis 55 81.81 37.78 0 14.28
Mandarin Citrus reticulata 50 86.00 27.91 9.68 0
Pomelo Citrus grandis 35 68.57 31.43 0 0
Lime
Figure 3.2. lotchy mottle in
leave
Figure 3.5. VLG’s sympto
ms in
orange fruit
Figure 3.6. Symptoms of
lotchy mottle and corky veins
Source (figure 3.2; 3.5 and 3.6): Lê Mai Nht, 2012
It can be concluded that when trees infected by VLG and tristeza in the same time
has forced the tree dieback very quickly, negatively damaging yield and quality of
harvested fruits.
3.1.6. Verification VLG’s infection ability in the field based on identical
symptoms
In addition to surveyed and collected samples, the study has applied
biotechnological method to verify the presence of virus and viroid in fruit trees (table
3.9). Result of this verification process using molecular method shows that tristeza
has occurred at all sample collecting sites. In Hanoi, presence of the three diseases:
tristeza, tatter leaf and exocortis in citrus crops. In Ngh An, exocortis has been
identified by RT-PCR and grafted into plant indicator (Etrog citron 861). In Bc
Quang, Hà Giang, samples of tristeza and tatter leaf diseases have been collected
(grafted into plant indicator Rusk citrange).
Table 3.9. Distribution of virus and viroid in some citrus fruit trees collected
from Northern provinces of Vietnam (Plant Protection Research Institute, 2011)
Location Crops
Citrus trees
Disease verification result
Tristeza
(Strip)

Hung et al., 1999 are very appropriate for a specific and in-depth study, genomes.
However, it needs a long time for DNA extraction using the above mentioned
method. For example, with 10 samples, the time needed for the whole process from
leave cleaning to DNA extraction is from 5 to 6 hours. It results in a high cost for
sample evaluation. Factors causing such a high cost include payment for labour
preparing sample, expenses on power and chemical. It is also noticeable that the use
of many toxic chemicals in this method has negatively impacted to human health and
polluted environment as well.
Such concerns have urged us to improve the method for DNA extraction by
using NaOH alternatively (table 3.10). This method is conducted in the National
Taiwan University of Science and Technology and in the same time, the method also
tested successfully in the laboratory of Plant Protection Research Institute.
Using NaOH to isolate DNA has saved time significantly, from 5 to 6 hours in
the old method to just from 45 minutes to 1 hour in this method, depending on skills
of technician. The advantage of this method is that DNA extraction is done by using
NaOH and acetic acid only, reducing considerably costs on labour and chemical
spending on DNA extraction process. The extracted DNA shows positive results in
VLG assessment.
Table 3.10. Improvement of DNA extraction method in diagnosing VLG (at the
laboratory in National Taiwan University of Science and Technology, 11/2011)
Tested
samples
DNA concentration collected in different DNA extraction methods
Regular extracted
solution
(Hung et al. 1999)
Pulverizing in
Eppendorf
by NaOH
Regular extracted

J
5
of
A. Hocquellet and Hung et al. (1999)] to assess VLG. Reaction of all primers have
shown an exact result in identifying causative agents of VLG. For citrus crops, this
method is a compulsory condition for trees to be formally certified as elite or disease-
free ones.
3.2.3. The density of psyllid Diaphorina citri Kuwayama on citrus plants
If fruit plants have been planted with fresh varieties without disease, the spread
of disease in nature will be vector D. citri Kuwayama. The diversity of varieties of
citrus as well as their different cultivation will be outbreeding faster and the disease is
able to adapt to server condition. Psyllar always occurred among 12 months in year
but they will have different densities. The densities of vector in citrus orchard depend
on food, cultivation and pest management, especially in relation to summer bud
season in year and season temperature. The densities of RCC are highest in summer
buds, spring season and decreasing in autumn and winter. In Nghe An, the densities
of psyllid is 25.15 individual/tree on non-cultivated orchards. And the densities of
RCC are very low on good-cultivated orchards (fig. 3.18). Figure. 3.18. The densities of Asian citrus psyllid on Xa Doai orchard in Nghe An
In 4 bud season in year, the densities of psyllid are highest in summer and
gradually descreasing in spring, autumn and winter. On non-intensive farming
orchard, the density of psyllid is 25.15 individuals/tree; in the meanwhile on the
intensive farming orchards only obtained 4.05 individuals/tree. Similar in spring bud
season were 21.5 vs. 3.7 individuals/tree, autumn season were 12.25 vs. 2.65
individuals/tree, winter season were 3.95 vs. 0.9 individuals/tree.
On Cam Canh orchard in Hanoi, the densities of psyllid were similar in Nghe
11


12

very susceptible to VLG with typical symptoms but this species is not belong to
genus Citrus; the results are consistent with Su et al. (In 2012). Similarly, the laurel
tree is allure tree for citrus psyllid as the domestic and foreign authors published.
Through the study, subjects initially detected and identified that Móc Mt tree
(Clausena excavata) is also one of good host plants citrus psyllid but not infected
VLG disease. So, it is maybe choice Tin Chót plants for breeding and mass
production of citrus psyllid, the laurel tree and Móc Mt tree is used to breed fresh
citrus psyllid resources citrus psyllid to design greenhouse experiments and extensive
experiments.
Table 3.17. The infected ability of Liberibacter asiaticus on some race of Rutacea
(Screen house Plant Protection Research Institute, 2011)
Species Vietnamese
Name
PCR’s diagnosis Rate
(%)
1
months
3
months
5
months
Citrus sinensis Cam ng ình - + + 100
Citrus unshiu Quýt ôn châu - + + 100
Citrus nobilis Cam sành - + + 100
Citrus reticulata quýt - + + 100
Citrus maxima i - - + 100
Citrus hyxtric Chp - + + 100
Citrus lemon Chanh - - + 100

Bacteria strain

Cam Xã
oà i
(Citrus
sinensis)

Cam
Canh
(Citrus
reticulata)

 i
Din
(Citrus
grandis)
Chanh
Eureka
(Citrus
limon)
Strain
I
Strain
II
1 G/x.-I-1-1 3 3 3 3 ×
2 G/x.-I-2-2 3 3 0 0 ×
3 G/x.-I-3-4 3 3 3 3 ×
4 G/x.-II-1-2 3 3 0 0 ×
5 G/x.- II -1 -3 3 3 2 2 ×
6 G/x.- II -1 -6 3 3 3 3 ×

16 G/x.-III-3- 4 2 2 0 0 ×
17

G/n.s
-
III
-
4
-

1

3

2

0

0

×18 G/n.s-III-4- 2 3 3 0 0 ×
19 G/n.s-III-5- 2 3 3 3 3 ×
20 G/c.l-VI-1-1 3 3 0 0 ×
21 G/c.l-VI-1-3 3 2 0 0 ×
22

G/c.


3

3

3×

Negative control 0 0 0 0 0 0
Total 14 10
14

Note: I: Hà Ni, II: Ngh An, III: Hòa Bình, VI: Qung Ninh; x.: cam Xã oài, n.s:
cam Nam Sn, c.l: cam lai, c.: cam ng, t.q: Orange China, Disease severity
was grade as follows: 0 = no symptom; 1 = mild chlorosis without dwarfing; 2 =
intermediate chlorotic mottling with moderate dwarfing; 3 = severe HLB symptom,
including yellow mottle, leaf hardening, and curling with distinct dwarfing

With 13 samples were collected from evaluated pomelo were obtained 7
samples infected with race I with and 6 samples infected with strain II. Strain II have
appeared in different varieties of grapefruit and also expressed different disease
severity levels of symptoms (table 3:20). Strain I and II cause disease on the citrus
varieties are summarized with symptoms in figure 3:25.
Table 3.20. The reaction of collected citrus plants from infected pamelo (Citrus grandis)
with greening disease (Screen house Plant protection research institute, 2013)

No


Strain

I
Strain
II
1

G/b.d
-
I
-
1
-
1

3

3

3

3×

2

G/b.d
-


0

0

×4

G/b.d
-
I
-
4
-
4

3

3

3

3×

5

3

3

0

0

×7

G/b.m
-
VI
-
3
-
4

3

3

2

2
-
5

3

3

3

3×

10

G/t.tr
-
VIII
-
1
-
6

3

3

3


VIII
-
3
-
1

3

3

3

3×

13

G/t.tr
-
VIII
-
4
-
1

3

3

technique, with two types of primer specificity (DNApol / nusG-rplK operon)
[Tomimura et al., 2010]. PCR products of primer DNA Poly obtained at 988bp; PCR
products of primer nusG-rplK operon obtained at 627bp. Two types of primer were
same reaction allows identification strains I, II and III. PCR products from the strains
I had 2 lines appear on products; II strains obtained product at 627 bp (figure 3:26).
Combining traditional methods with molecular biology have identified pathogenic
strains greening disease in Vietnam and the distribution of pathogenic bacteria in the
survey area (figure 3:27).
Figures.

3.25. Strain I cause disease on Cam Xã
oài and Cam Canh;
Strain II cause disease on 4 varieties (bi Din,
chanh Eureka, Xã oài và cam Canh)

Figures.

3.26. The duplex
-
PCR results and strain
identification [CC: Cam canh, RL=Rumprue
lime, Cal=calamondin (Qut)].
Strain I: obtained PC R product at 988bp and
627bp; Strain II: obtained PCR product at 627bp
(Sources: Lê Mai Nht, 2013)
with Trng Vng orange variety (Plant Protection Research Institute, 2010)
Rootstock age
Success rate (%) Un-success rate for
died bud (%)
Un-success rate for
died callus (%)
1 week-old 10.0
c
10.0
c
80.0
a

2 week-old 34.0
a
26.0
bc
40.0
b

3 week-old 28.0
ab
38.0
b
36.0
bc

4 week-old 20.0
b
70.0

MS + 0,5mg/l BAP 44.3
a
5.4
a
9.8
a

MS + 1,0mg/l BAP 39.4
ab
4.2
b
7.0
ab

MS + 1,5mg/l BAP 35.6
b
3.6
bc
3.9
b

CV(%) 12.3 19.7 21.1
In order to improve shoot tip grafting, the project were optimaled technique in
apical grafting. The result is optimal in the development of research methods of Su
(2008), applied in the laboratory of the Plant Protection Research Institute and
17

National University laboratory synthesis Taiwan. The media was used in liquid MS
medium supplemented growth regulator (i-inositol, thiamine-HCl, pyridoxine-HCl,
nicotine).

micro-
grafting
(%)
Ratio of
un-
infected
greening
disease
(%)
Ratio of
un-
infected
tristeza
disease
(%)
Meristem Phc Hòa Pumelo 30 0 - -
Trng Vng
Orange
30 0 - -
Meristem with
two leaf
primordia
i Phc Hòa 30 26.7 100 63.3
Cam Trng Vng 30
33.3
100 73.3
Meristem with
three or four
leaf primordia
i Phc Hòa 30 36.7 73.3 23.3

-
old

13

15.
4

16

18.
8

6 week-old 14 78.6 13 76.9
8 week-old 10 90.0 12 91.7
In micro- apical grafting technique, bud stage processing is concerned, used
buds in grafting were very vulnerable. Through practical manipulations buds, we had
tried to find solutions to overcome this phenomenon. Results were determined by
using 2-3 drops of Tween 20 for in sanitizing solution and sterile distilled water.
When the buds are located near the lower part suspended in the cup containing the
disinfectant solution, remove the disinfectant solution and rinsed with sterile distilled
water, the buds put into again in the cup, easy operation, bud not damaged, crushed.
Tween 20 also had the effect of seepage slicks in the armpit leaves sprout, so better
steriled effects and incidence of less fungal contamination.
3.4.2. Production of disease-free citrus seedlings in 3-level grid system greenhouse
Seedling quality is an important factor in developing citrus fruit. To ensure
disease-free planting must be produced in the 3-level grid system greenhouse and
resources of grafts were taken from the apical meristems of grafting and has been
tested and recognized disease-free certification.
Improved potting mix with ratio fifth yellow sand + 2/5 wood shavings + 2/5

rootstock), 33.33% (Chp rootstock).
Table 3.35: The percentage of disease after artificial infection in infected grafts
for rootstock is used in seedling production
(Screen house Plant protection research institute, 2012)
Rootstock Resources
(infected
grafts)
After 2 months After 4 months After 6 months
TLB
(%)
Disease
level
TLB
(%)
Disease
level
TLB
(%)
Disease
level
Sour Pomelo Strain I 0.0
c

0 28.89
c

1 51.48
c

3

LSD
0.05

0.
946.
047.
32CV(%) 3.4 5.9 4.8
Note: Strain I: from bi Din; Strain II: from cam Nam Sn
Pathogens of greening disease had direct influence to rootstock, which made
limit grow height of rootstocks. During follow-up, potentially pathogenic strains
mutated virulent infection in the next generation. The same strain of the disease on
the same object in both rootstocks but obtained was lower plants. The phenomenon of
dwarf plants has demonstrated symptoms of low dwarf tree in the field. This result is
consistent with studies of Su et al., (2008, 2012); (Hung et al., 2012) to assess
tolerance of grapefruit and Chp.
20

3.4.3.2. Tolerance of imported rootstock to greening disease
The imported rootstock from Taiwan (Cleopatra mandarine, Swingle
citrumelo, Fruit dragon, Volkamer lemon, Poncirus trifoliata, Carrizo citrange,
Troyer citrange, Rangpur lime) are grown and developed in Vietnam to find out the

8 Rangpur lime

2 3
9 Chp 2 3
10 Bi chua 1 3
Note: Infected grafts from cam Nam Sn; - : 0 = No symptom (tolerant), 1 = Mild (blotchy
mottling symptoms observed from 1 to 30% on seedlings canopy, 2 = Moderate (yellowing
symptoms observed from 31 to 50% on seedlings canopy), 3 = Severe (blotchy mottling, midrib
yellowing and twigs dieback symptoms observed more than 50% on seedlings canopy).
3.4.4. Against re-infected greening disease in the fields
3.4.4.1. Using antibiotics for control infected grafts
The previous results always used Tetracyline 1% for control greening disease
(Aubert và Bové, 1980; Tsai et al., 2012). In this research, based on mechanism of
pest, antibiotics in control infected grafts, is Streptomycine (suitable for negative
gram bacteria)
Table 3.38. Effect of Streptomycine to greening disease symptom
(Screen house Plant Protection Research Institute, 2012)
Experiment Total
plants
Survival
rate (%)
Symptom after infection
2
months
4
months
6
months
Steptomycine 1% + 12 hour 20 100 + + +
Streptomicine 1% + 24 hour 20 90 - - -


Free disease seedlings + pesticide spraying
frequently
0.0
a
1.33
a
5.33
a
Free disease seedlings + local cultivation 4.0
b

10.67
b

16.0
b

LSD
0.5
0.66 1.15 1.99
CV (%) 7.5 6.5 9.4
Note: initial evaluation disease rate after 15 months planted; planted: 25/12/2011; 1
st

evaluation: 15/3/2013; 2th evaluation: 15/7/2013; 3
rd
evaluation: 15/10/2013
After 15 months planted, re-infected plants occurred in “no pesticide spraying”
method for control vector (9.33%), in “local cultivation” (4%). In “pesticide spraying

diseases. Should be planted legumes to improve soil.
- Planting windbreaks fence around orchards with acacia.
- Check the pH of the soil in each region to have appropriate
rehabilitation measures
Seasonal Should be planted in spring, autum
Densities of
planting
- Orange variety: 650 – 750 plants/ha
- Mandarine variety: 800 – 1000 plants/ha
- Grapefruit variety: 450 – 500 plants/ha
Size hole for
planting
- Mountainous: 1m × 1m × 1m
- Flat, well drainage: 0,8m × 0,8m × 0,8m
- Land was transformed from rice, vegetable, need to make bed
and upper planted to avoid plant flooded.
Cultivation - Weeding (hoeing grass around the base of the foliage, cut
foliage grass outside)
- Cover the dried straw mulching
- Fertilization: Basic construction period applied 3-4 times /year;
trading period applied 2-3 times/year
- Watering: centralized irrigation in the dry season
- Pruning, Canopy
Chemical Control Psyllar vector in new buds sprout stage by using Confidor
100SL or autoclaves or systemic absorption (if any vector)
Training - Identify vector and major pest
- Follow the rules and characteristics arising pest
- Integrated management of pests

The effects of macro elements and trace: N: increased growth potential and

use toxic chemicals.
The VLG disease showed rapid symptoms on Xã oài orange, Cam Canh after
45 days of grafting. The density of RCC was 6 individuals/tree or 9 individuals/tree
in 24-hour exposure, symptoms (respectively) appears only after 9 months and 6
months of infection on Xã oài oranges and Cam Canh.
It is the first time to identify Tin Chót (Severinia buxifolia), which was
collected in Tho Xuan, Thanh Hoa, is host with greening disease.
1.3. By the reaction of VLG samples for bio-monitoring plants and a number of
fruit trees (orange, tangerine, grapefruit, and kumquat) combined with duplex-PCR
techniques have identified bacteria strains in the North belonging to VLG strain I and