Occurrence of plant bacterial diseases in Jordan

Different studies were done in order to investigate the occurrence of bacterial diseases on different crops grown in Jordan during different growing seasons. Surveys were made and different bacterial diseases have been recorded based on symptoms and pathogenic nature. Morphological characters, biochemical tests and PCR detections were employed in order to detect and identify the causal agents of different inspected plant bacterial diseases. In addition, the distribution of the identified bacterial diseases, throughout the country was recorded. The results of our study revealed the occurrence of different bacterial diseases attacking different crops; grown in many growing regions throughout the country. Some of them were found to have a wide host range such as crown gall and soft rot, while others had a restricted host range as in the case of bacterial speck of tomato which was found to be restricted to tomato and black leg of potato. As a result of this study, the following diseases; angular leaf spot of cucumber, tomato speck, common blight, crown gall, soft rot, black rot, black leg and bacterial cankers resulted in high economic losses in yield. The spread of these diseases in the different areas in Jordan with different environmental conditions may result in the development of new races of the causal agents without developing typical symptoms making their diagnoses under field conditions difficult. Whereas the bacterial diseases needs deep and ideal studies in order to diagnose diseases, the diagnoses of these diseases act as the base for researchers to challenge and withdraw researches into the improvement of novel, more effective and sustainable bacterial disease control strategies.


INTRODUCTION
The world's population is increasing every year and in order to meet their demands, global crop production needs to be increased.Plant diseases attack all agricultural crops which are considered as the main source for human food and clothing all over the world and considered among the main factors that drastically affect its production, resulting in economic losses either in the field or in storage by decreasing crop production in quality and quantity.Thus, one of the methods to increase food production is to control plant diseases.Plant disease causal agents could be; fungi, bacteria, viruses and nematodes.Different bacterial diseases have been reported to attack many agricultural crops around the world, leading to high economic losses in yield under favorable environmental conditions and could eliminate plantation of certain crops in certain areas as in the case of fire blight (Erwinia amylovora) attacking pome fruits in northern states of United States of America (Kennedy and Alcorn, 1980;Agrios, 2005).
Moreover, plant bacterial disease diagnoses are very difficult under unfavorable conditions were typical symptoms do not develop and could be masked with other disease symptoms as in the case of black rot of crucifers caused by Xanthomonas campestris pv.campestris; a humid, warm season pathogen while crucifers hosts are winter season crops, typical symptoms will not develop under cold conditions (Mahiar and Khlaif, 2000).
Plant bacterial diseases spread in tropical and subtropical regions, and host range of bacterial diseases varies according to the disease, however some of them are with a wide host rang such as soft rot Pectobacterium carotovorum subsp.carotovorum (Omar andKhlaif, 2000 andAbu-Obeid et al., 2017), and crown gall; Agrobacterium tumefaciens (Al-Karablieh and Khlaif, 2002;Al-Karablieh et al., 2006).On the other hand, some bacterial diseases had a narrow and restricted host ranges as in the case of black leg of potato P. carotovorum subsp.atroseptica (Al Masa'adeh and Khlaif, 2003;Masa'adeh and Khlaif, 2004;Abu-Obeid et al., 2017) and bacterial speck of tomato Pseudomonas syringae pv.tomato (Abu-El Samen and Khlaif, 1999).
Recently the interests in bacterial diseases have been increased due to its importance and its serious damage on different crops resulting in great losses in quality and quantity.

Economic importance
Losses due to bacterial diseases are difficult to estimate, especially in fruit trees where losses are not confined to the year of disease development.
The prevalence of favorable environmental conditions leads to high economic losses to the crop as in the case of angular leaf spot of cucumber under plastic houses (Khlaif, 1991a and b).
Ralstonia solanacearum (Pseudomonas solanacearum); the causal agent of vascular wilt disease of solanaceae, ranked the second most important bacterial pathogen (Mansfield et al. 2012).The pathogen is distributed worldwide and induces a destructive economic impact.Direct yield losses by R. solanacearum vary widely according to the host, cultivar, climate, soil type, cropping pattern, and pathogen strain.Due to R. solanacearum, yield losses were estimated from 0 to 91% in tomato, 33 to 90% in potato, 10 to 30% in tobacco, and up to 80 to 100% in banana in Africa, India and Indonesia (Kelman, 1998).
Xanthomonas oryzae pv.oryzae causing leaf blight of rice is a major disease of rice and is a serious threat to rice production in both temperate and tropical rice-growing regions, due to its high epidemic potential causing up to 60% loss in India and Indonesia (Ou, 1985;Raina et al., 1982).
Canker and gummosis of stone fruit trees caused by Pseudomonas syringae pv.syringae, are of major concern in fruit producing areas worldwide, and are exceedingly and difficult to control, and resulted in significant economic losses.In 1967 about 750000 peach trees were killed in France and resulted in 10-75% losses of trees and reduction in yield reached 10-20 % (Luisetti et al., 1976).
In Florida citrus canker, caused by Xanthomonas citri subsp.citri, led to the destruction of half million citrus seedlings and bearing trees and to millions of dollar losses (Kennedy andAlcorn, 1980 andKelman, 1998).
Bacterial blight of beans causes reduction in yield and decreases the marketable value of the crop, including seed size, quality and may range from a trace to 100%, especially when favorable environmental conditions persist during the early growth and flowering stages.Kennedy and Alcorn (1980), estimated losses in dry edible beans due to bacterial blights which ranged from 75 to 90% in North Dakota, while in Jordan Valley losses of bean yield due to common blight estimated ranged from 73 to 85% of the total production (Khlaif and Qadous, 1995).
Production of tomato seedlings free from bacterial speck is impossible, especially under plastic houses conditions in the Jordan Valley; 75% of tomato fruits showed specks which resulted in a 22% yield reduction (Abu-El Samen and Khlaif, 1999).
Angular leaf spot disease of cucumber, caused by Pseudomonas syringae pv.lachrymans was found to reduce the yield of cucumber at a rate of 30.6 to 64.4% in the Jordan valley and about 50-93% in the Uplands of Jordan (Khlaif, 1991a,b,c;Khlaif and Abu-Blan, 1994).
Due to lack of information about the situation of bacterial diseases in Jordan, this study was undertaken in order to inspect, diagnose, isolate, and identify the causal agents of bacterial plant diseases and their occurrence in Jordan over a period of more than 15 years.

MATERIALS AND METHODS
Extensive field surveys were undertaken to inspect, detect the occurrence, prevalence and the incidence of different bacterial diseases affecting economic crops, including; vegetables, fruit trees and ornamentals planted in different agricultural areas in Jordan.Through field trips to different agricultural areas during the different growing seasons in Jordan, samples of different crops suspected to be infected with different bacterial diseases throughout different growing seasons in Jordan, were collected, and placed in ice box and brought to the laboratory for further identification.

Isolation and identification of the pathogenic bacteria
Initial identification was performed according to Schaad et al. (2005), through isolation by using differential, common and semi selective media as recommended for the suspected bacterial pathogen.A small amount of suspected tissue was removed from plant parts suspected to be infected with a certain bacterial disease, with sterile scalpel, washed and rinsed with sterile distilled water (SDW) and disinfected with diluted bleach solution (0.5% sodium hypochlorite), after rinsing in sterile water, tissue was chopped up with a sterile scalpel in a droplet of SDW a n d l e f t t o stand for 15 min.The bits of surface sterilized tissues were transferred onto the surface of loop and the resulted suspension was streaked on the surface of dried media plates.The media used for isolation were selective media according to the suspected pathogen as for example; D1 media was used in the case of crown gall (Fakhouri and Khlaif, 1996;Al-Karablieh and Khlaif, 2002;Al-karabliah et al., 2006), BCBRVB in the case of fluorescent Pseudomonas (Abu-El Samen and Khlaif, 1999;Hijazin and Khlaif, 2005), Logan media in the case of Pectobacterium (Masa'adeh and Khlaif, 2004;Abu-Obeid et al., 2017), TZC in the case of Ralstonia solanacearum, NA and SX Agar in the case of the Xanthomonas campestris pv.campestris (Mahiar and Khlaif, 2000), on the other hand, Nutrient Agar and KB media were used as common media for other pathogens.Inoculated media plats were incubated at 25 ± 1°C, checked periodically for development of suspected phytopathogenic bacterial colonies, and then subculturing was made by transferring a suspected colony to a new media plate through streaking for purification.Then, the obtained purified suspected colonies were transferred into agar slants and allowed to grow, kept in refrigerator for further identification.
Then the colonies of the suspected bacterial pathogen causal agents were subjected to identification procedures through biochemical and physiological tests as recommended by Schaad et al. (2005).The same tests were run against a reference culture of the most different identified pathogens.

RESULTS AND DISCUSSION
Approximately, all growing regions planted with different crops have been inspected and different bacterial diseases have been recorded in cultivated and noncultivated crop plants of Jordan.The results of this study indicated the occurrence of different bacterial diseases in the different inspected agricultural areas throughout the country.The different diagnosed and identified bacterial diseases and their causal agents are listed in Table 1.
However, the symptoms of the diagnosed and identified bacterial diseases vary from leaf spots, tumors, rots, wilts, scab, cankers, and gummosis etc., indicated by different causal agents.
The occurrence of these diseases depends on the geographical areas since Jordan is divided into different geographical areas of Jordan Valley, Jordan Valley rift, mountains or uplands and deserts (Figure 1) where the environmental conditions of these areas varies according to the season and location.In summer, it is very hot and dry in the desert, hot and dry in Jordan Valley, while it is warm and relatively humid in the uplands and mountains.In winter, it is warm and humid in Jordan Valley, cold and dry in the desert, at the same time cool and humid in the uplands.These wide climatic variations in Jordan could illustrate the importance of different plant bacterial diseases in relation to occurrence, development and spreading, as well as, under certain conditions could lead to an epidemic bacterial disease.
However, eight phytopathogenic bacterial genera were identified; Agrobacterium with two species were recorded.However, A. tumefaciens was found to be the most common, causing crown gall attacking 21 different hosts, including; stone fruits followed by grapes, roses, olives, quince, pomegranate and nemaguard.The disease was found to spread throughout the country in different climatic regions.Also, A. rhizogenes was with two hosts causing hairy root disease on apples and roses.
Erwinia with seven species, causing rots and blights were identified and recorded, from which the species Carotovorum with had two subspecies; Carotovorum and Atroseptica; Erwinia carotovra subsp.cartovra was found with a wide host range of about 30 different host plant including; vegetables and ornamentals and was found to spread all over agricultural regions either in field and storage (Khlaif, 1993).On the other hand, the subspecies Atroseptica has been found to be restricted to potato, causing black leg disease in winter in the Jordan Valley region and early spring in the uplands.
Erwinia amylovora the causal agent of fire blight disease on pome fruits attack four hosts; apple, pear, quince and firethorn in spring where it was severe on the flowering stage and was more serious on pears and apple (Al-Dahmashi and Khlaif, 2004).Other species of Erwinia such as E. chrysanthmi (Dickey dadantii), E. trachephila and E. stewartii were identified causing wilt, rot and leaf blight diseases on different hosts in many agricultural regions of Jordan (Table 1).
Seventeen species of Xanthomonas (Table 1) were identified and detected on different types of hosts, where X. campestris pv.campestris was the most common and reported on 10 different hosts causing symptoms ranging from black rot on crucifers to leaf spots and blights on legumes.Also, it was isolated from a wide range of weeds spreading in the same regions or fields of infected crop.However, different phytopathogenic bacteria genera such as X. campestris pv.phaseoli and Xanthomonas campestris pv.campestris were isolated from different weeds and volunteer plants grown in the host fields (Table 1).These finding could bring a new dimensions in the epidemiology of these diseases and play an important role as a source of inoculums for these diseases.
X. arboricola pv.pruni attacked 6 different hosts of stone fruits causing leaf spots where it infects more seriously peaches, apricots and plums, especially in the  1).Different species of Pseudomonas were isolated and identified, of these P. syringae pv.syringae was the most common species causing many diseases on 14 different hosts; brown spot, scab, wilt, cankers, citrus blast and bud blast.Other Pseudomonas species attacked 24 different hosts causing different disease symptoms ranging from angular leaf spot such as ALS (P.syringae pv.lachrymans) on cucurbits especially under plastic houses, gummosis (P.syringae pv.morsprunorum) on stone fruits, knots (P.savastanoi pv.savastanoi) on olives, oleanders, jasmine and Ziziphus sp. and spread all over the country, causing a serious damage to all olive cultivars; Nabali baladi was found to be the less olive cultivar susceptible to olive knot disease (Khlaif, 2006).
Streptomyces scabies was found to cause common scab disease on potatoes, carrot and radish and was most common on potatoes in the southern parts of Jordan where the soil is sandy.
Ralstonia solanacearum was isolated and identified from wilted plants of tomato under plastic houses and banana in the Jordan Valley during spring.
One species of Clavibacter (Corynebacterium) was detected and identified, Clavibacter michiganensis pv.michiganensis causing Bird's eye spot disease on tomato was recorded during winter in the Jordan Valley.

Conclusion
Many plant bacterial diseases were detected, identified and recorded that cause diseases of many crops grown in Jordan which resulted in high economic loss.Accordingly, the identification of these diseases using efficient methods is essential in order to investigate the ecology of these diseases and therefore help in employing the efficient control method.The occurrence of these bacterial diseases in the different growing areas in Jordan with different environmental conditions could develop new phytopathogenic bacterial strains, where atypical symptoms could be very difficult to diagnose and control.
This work acts as the base for researchers in this field to take advantage and implement further studies in controlling these diseases and decrease losses.

Figure 1 .
Figure 1.Map of Jordan showing the main physiographic regions.Source: Atlas of Jordan, 2014.

Table 1 .
List of diagnosed and identified plant bacterial diseases and their causal agents in Jordan.
campestris pv.phaseoli causing common blights and leaf spots of bean and peas.On the other hand, X. campestris pv.translucence was isolated and identified from 3 field crops showing strips or streaks symptoms; the disease was found to be common during winter and spring in the Jordan Valley and in the uplands, where barley was found to be the most susceptible crop.Other 13 Xanthomonas species with narrow host range, attacking different plant types ranging from ornamentals, fruit trees and vegetables were diagnosed and recorded (Table