Incidence of Cereal Cyst Nematode Prevailing in Wheat-rice Growing Regions of Punjab Province, Pakistan

Cereals are the most important food source in the world and 58 per cent of the annual cultivation has been allocated to wheat, corn and rice. By the year 2030, the world population will reach about 8 billion people and grain consumption will be increased significantly (Curtis, 2002). Among cereals, wheat and rice are the main cereals grown in Pakistan. During 2017-18 marketing year, wheat production was estimated as 19.1 million metric tons with an area of cultivation of 6.56 million hectare. The rice is the third largest crop in terms of area sown, after the wheat and cotton crops. Rice production in 2017-18 marketing year was estimated as 3.89 million metric tons with an area of 1.85 million hectare (Anonymous, 2017-18). The production of wheat and rice is expected to increase in coming years.
 
Cereal cyst nematodes (CCNs) are known as the utmost vital nematode pathogens that reduce the growth of cereal crops. Cereal cyst nematode parasitize the newly developing roots of plants and reduce the crop vigor by producing patches of stunted plants with pale green leaves. Young plants develop less number of tillers and root system turns shallow and have a “bushy-knotted” look (Nicol et al., 2010; Smiley and Nicol, 2009). Cereal Cyst nematodes (CCNs) of genus Heterodera include approximately 70 species, having a multipart of 12 species known as from the group of Heterodera avenae (Rumpenhorst et al., 2003). Twelve species from Heterodera avenae species complex viz., H. australis, H. filipjevi, H. mani, H. pratensis, H. riparia, H. sturhani and H. ustinov, H. arenaria, H. latipons, H. avenae, H. mothi and H. hordecalis have been characterized by Subbotin et al., (2001).  Maqbool (1981) noticed dominance of cereal cyst nematodes such as H. avenae, H. zeae and H. mothi in the soil samples of wheat fields. Maqbool and Kerry (1997) confirmed the presence of cereal cyst nematodes and identified six new cyst nematode species associated with grain crops in Pakistan. During 2003-2004, Shahina and Erum, (2007) conducted a survey in the southern regions of Pakistan and found that 39% of total studied area was infected by cereal cyst nematodes through huge dispersal and host choice of corn cyst nematode (H. zeae). The cereal cyst nematodes have shown significant yield losses in specific research trials by as much as 50% in Australia, 20% in Pakistan, 90% in Saudi Arabia and 50% in Turkey (Riley et al., 2009). More than half of the growing areas of cereals are detected to be infected by CCNs in certain cereal growing areas of Turkey (Abidou et al., 2005; Rumpenhorst et al., 1996), Europe (Cook, 1974) and USA (Smiley et al., 2009). It is also observed that approximately most of the wheat fields are infected by cereal cyst nematodes in China (Peng et al., 2010).

The present study was conducted by keeping in view the sparse data on distribution of cereal cyst nematodes, their population densities and most abundant species in specific cereal growing regions. During the surveys, samples were collected from the wheat-rice monoculture fields of Punjab province, Pakistan. The study provided comprehensive information on cereal cyst nematodes population and species.

No. of nematodes in 1 ml suspension × Total volume of suspension.

 
 

Population densities of CCNs in wheat and rice growing regions
 
Cereal cyst nematodes (CCNs) were found in 80% of wheat and rice fields surveyed. The results showed that in 160 soil samples collected from wheat fields, 20 samples (12.5%) were noticed to have infection. The number of cysts in wheat fields were varied from 7-38 cysts/100g soil, the eggs and juveniles varied from 142-771 (Table 2). Out of 90 rice fields sampled during the survey, on average 22.5 (25%) samples were found infested with CCNs. Population density of CCNs was varied from 17 to 25 cysts/100g of soil with 345-508 of eggs and juveniles. The results revealed that cereal cyst nematodes are present in all the localities surveyed viz., Jhang, Khushab, layyah, Hafizabad, Sheikhupura, Narowal, Gujranwala and Sargodha with abundant population densities.
 

 
Prevalence of cereal cyst nematodes across regions
 
The prevalence of cereal cyst nematodes (CCNs) were assessed from all wheat-rice growing regions surveyed.  In the wheat fields, 6.25% and 16.88% are the lowest and  highest incidence of CCNs recorded respectively in Sargodha district. In rice fields, 24.69% to 27% incidence was recorded in Gujranwala and Hafizabad respectively. The average numbers of cysts, eggs and juveniles per 100 g of soil in wheat fields was higher than in rice fields (Table 2). The presence of abundant cysts with healthy eggs and juveniles were observed in most of the samples (Fig 2).  Moreover, the aged fields were found to be highly infected with CCNs (Fig 3). The results showed that in Punjab province, nematode population density index indicates that about half of the wheat and rice fields are moderate to highly infected and proper management practices need to be adopted.
 

 

 
Morphological identification of cereal cyst nematodes
 
The morphological based identification of CCN species was conducted by picking the second stage juveniles from the hatched cysts. The results showed the presence of three Heterodera species in all the samples collected from rice and wheat fields. The Heterodera oryzae was more abundant than other species in rice fields and Heterodera avenae was found more abundant in wheat fields. Heterodera graminophila was present in both rice and wheat fields with other two species. Heterodera oryzae is certainly the only species of Heterodera to keep cysts with cone vulvae and juvenile in which the hyline portion of the tail is clearly longer than the stylet (Fig 4). While, cysts are lemon shape, dark brown to black, whose dimension was ranged from 0.31-0.81×0.22-0.69mm (average 0.57×0.45mm), with having zig-zag ornamentation without order defined and partially covered by semi-crystalline layer. The length of second stage juvenile (J2) ranges from 0.37 to 0.50mm with an average of 0.44mm long. The second stage juveniles (J2) of Heterodera avenae are vermiform and cylindrical with a sharp pointed tail as shown in (Fig 4). Stylet shape is slender and strong with large anteriorly flattened to concave basal knobs. Head section is hemispherical and offset. Middle bulb is round with a large valvuae apparatus inhabiting about half of the corresponding diameter of body (Fig 4). Tail is 3-4 times anal body thickness long. Tail is conoid, gradually narrowing to a finely rounded station. The body of Heterodera graminophila was observed as pearly white, usually rounded shaped, with protruding vulva and neck. Thick cuticle with zig-zag pattern, subcuticular punctation and usually a granular surface obscuring details of pattern. H. graminophila have strong stylet and curving dorsally, with sharp-developed knobs sloping posteriorly. Moreover, esophageal part appearing about as illustrated, with excretory pore normally at base of neck. Small anus, inconspicuous and located about 20% of body length from vulva (Fig 4). Cyst surface in H. graminophila usually granular, with conspicuous irregularly arranged function. The morphological observations revealed the presence of major Heterodera species in all the rice and wheat cropping fields.
 

 
The present study of cereal cyst nematode (CCN) revealed the incidence of cereal cyst nematode in different rice and wheat growing regions of Punjab province Pakistan. Some fields surveyed were found to be highly infected with CCNs while some fields recorded less infection. Number of cyst and nematodes per 100gm of soil were varied in different regions ranging from 17-38 cysts and hundreds of eggs and juveniles. Ahmadil et al., (2014) reported varied degree of cereal cyst nematodes across the different regions of Iran. The variation in the infestation of CCNs occurs due to the different factors like application of nematicides, soil solarization, cultural practices or the presence of fungal and bacterial pathogens (Chen, 2007). Cereal cyst nematodes have more devastating effect on rain-fed cereal crops than irrigated crops (Smiley and Nicol, 2009).

According to Rivoal and Cook, (1993), now a days, the use of resistant or tolerant varieties to the nematodes is considered as one of the most efficient management strategies for controlling the CCNs and is broadly used in some countries such as Australia, Denmark, Sweden, England and France. However, during the survey, the CCNs were observed in different local varieties of wheat and rice under field conditions, it would be helpful to examine reaction of a wider range of wheat and rice cultivars under controlled conditions. CCNs population was found high in wheat fields than the rice fields. Andersson (1982), reported that spring wheat is a more suitable host for H. avenae.

The given results showed the trend of increasing population densities of CCNs with respect to years. The level of population was found to be related with the age of fields. Aged fields were found to have more population density of CCNs. The study showed that crop rotation and fallow effects the population of cereal cyst nematode in wheat crop (Taya et al., 2000). Three CCNs species Hetrodera avenae, Heterodera oryzae and Heterodera graminophila were found in these cereal growing regions. The microscopic observations of anatomical features of these nematodes described the specialized body parts in all species. The observations of cysts were also made and vulva slit on cyst approximately about 45µm in length (Kumari, 2017). The morphological observations of cereal cyst nematodes were clearly demonstrated by Golden and Bircfield, (1972). The number of J2 and eggs of CCNs in some regions were greater than that measured as damage threshold level for this nematode (Gill and Swarup, 1971; Meagher and Brown, 1974) and is expected that these populations could cause economic yield loss. The most effective control method for CCNs is crop rotation with non-host crops. Some other control methods includes clean fallow and deep ploughing of 2-5 times during May to June in India (Swarup and Sosa-Moss, 1990), also early planting of wheat in order to increase the plant tolerance and vigor against nematode attack and application of nematicides in planting (Brown and Kerry, 1987). This study will provide a comprehensive data on cereal cyst nematode population and distribution across wheat-rice growing regions of Punjab province of Pakistan and development of resistant cultivars and management strategies can be designed to improve the yield of these both important cereal crops.  

The soil samples were collected from wheat-rice growing regions of Punjab province of Pakistan to study incidence of cereal cyst nematodes (CCNs). The incidence of 6.25% to 16.88% was found in wheat fields. In rice growing areas, the incidence was 24.69% to 27.00%. Through this study, incidence and prevalence of Heterodera spp. were recorded for the first time from wheat- rice growing regions of Punjab province, Pakistan. 

The funding was provided by University of Sargodha under ORIC-UOS projects (UOS/ORIC/2016/16).