Quantifing Diversity and Phenology of Cereal Leaf Beetles Oulema Spp. (Chrysomelidae: Criocerinae) in Several Cereal Fields of Semi-Arid Zone, Algeria

The present study treated the taxonomic diversity of the cereal leaf beetle (Oulema spp.) community subservient to several cereal fields (barley, durum wheat and oats) in Eastern Algeria. From April to June, which is the appearance period of these insects, the leaf beetle fauna was weekly sampled using two trapping techniques: entomological net and sight hunting method. Spatiotemporal alpha and beta diversity of this community were evaluated by several ecological indices: Shannon index, Simpson index and equitability. The multivariate statistic test (GLM) was used to assess the variation of the different diversity parameters. Catches totaled 1144 individuals belonging to the genus Oulema and four species: Oulema melanopus, Oulema cyanella, Oulema duftscmidi and Oulema gallaeciana. The highest abundance of the leaf beetles was recorded in oats with 569 individuals (49.73%), followed by barley (390 individuals, 34.09%) and durum wheat (185 individuals, 16.17%), whereas the most speciose field was barley (4 species), followed by oats (3 species) and durum wheat (2 species). The most abundant species was O. melanopus with 746 individuals (65.20% of the total). The spectrum of occurrence ranked constant species first, followed by common and very accidental species. GLMs demonstrated that diversity of cereal leaf beetles Oulema spp is highly related to cereal field type.


INTRODUCTION
Often called leaf beetles and with about 19 subfamilies and 37 000 known species, Chrysomelidae is considered as a very diverse family among insects that feed on plants (Ekiz et al., 2013). Specifically, according to Bezdĕk and Schmitt (2017), the subfamily Criocerinae contains 1500 species devided into 22 genera in the world, of which about 200 are found in the Palaearctic countries. The larger part of the species lives in subtropical and tropical regions, and belongs to five genera: Crioceris, Lilioceris, Lema, Oulema, and Neolema (Matsumura et al., 2014). The genus Oulema comprises 800 descriped species, a variety of them are serious cereal pests like O. gallaeciana (Ulrich et al, 2004) and O. melanopus, which is the most dangerous (Vencl and Leschen, 2014). Indeed, the larval and the adult forms of this taxonomic group prefer living and feeding on leaves. Hence, a lot of species attract many researcher's attention from different specialities for their feature as phytophagous (Booth et al., 1990) and their aptitude to cause considerable damage to some crops, especially cereals (Weloso, 1973). In Algeria, products of cereals have an important position in food system and in economy of the country; this characteristic is clearly perceived through all the phases of the sector (Djermoun, 2009). The great fluctuation of yields and production of cereals are in large proportion the result of abiotic constraints such as thermal and water stress (Derbal et al., 2015), and biotics such as pests (Belahcene et al., 2015).
The Algerian entomofauna restricted to cereal fields is not well assessed. In fact, few works have been conducted in this context, e.g: Madaci (1991) in El khroub, Kellil (2010) in Setif, Bakroune et al., (2020) in Biskra, Amokrane et al., (2020) in Oum El Bouaghi. As far as the genus Oulema is concerned, it remains poorly studied, despite the potential danger that some of its species represent for cereal production. According to Sahraoui et al. (2001), the species O. melanopus is the main cereal pest in Algeria. To our knowledge, the notable works dedicated to this genus are Rouag et al., (2012) and Kellil et al., (2018), the first evaluated the infestation by Oulema spp. on six varieties of durum wheat in arid conditions of Setif, and the second studied the population dynamics of O. melanopus in different durum wheat varaities of the high plains of Eastern Algeria.
The basic knowledge on species and their distribution are of paramount importance for the biodiversity study in a given region (Sarthou et al., 2005), particularly for those living in hard environments like arid and semi-arid areas (Chenchouni, 2012). Consequently, this study, which is the first in Algeria to pay particular attention to the diversity of cereal leaf beetle community (Oulema spp), attempted to collect data on this community in many cereal fields located at semi-arid zones. Many research questions could be asked in this context: (i) under semi-arid climatic conditions, what would be the distribution pattern and the structure of cereal leaf beetle Oulema spp. communities in different cereal fields? (ii) does the spatiotemporal variation in species richness and abundance exist?

Study Area
This survey was carried out in two regions located in Eastern Algeria ( Figure 1) The annual average of mean temperature was 16.62 °C with a maximum in July (27.50 °C) and minimum in January (7.60 °C) in Tebessa, nevertheless, in Batna, the annual average of mean temperature was 15.82 °C, the maximum temperature was recorded in July with 28.79 °C, and the minimum was recorded in January with 4.70 °C.

Sampling and Identification
Fieldwork was performed weekly and regularly in the three cereal fields (12 outputs per field), from April, the date of appearance of the first adult, until late June, the harvest day, during the year 2011.
Since adult leaf beetles are generally found on the upper leaves of wheat, two trapping techniques were adopted: (i) trapping by using an entomological net (35 cm of diameter), (ii) the sight hunting method, in order to complemente the first technique. Every field was divided into 10 plots, in each plot we gave 50 shots randomly, progressing in the direction of the field length. The sampling session took over three hours from 10:00 to 13:00 AM. In the laboratory, the identification of the collected biological material (adults) was performed by using mainly these keys: Lacordaire (1845) and Perrier (1971).

Data Analysis
The diversity of the collected leaf beetle community was evaluated using different parameters: (i) relative abundance (RA) which is the number of individuals of a species / the total number of individuals (N), (ii) average of abundance (AA) represents the number of specimens of a given species (Magurran, 2004). Occurrence frequency (Occ) which is the number of times the species was detected on the number of all samples. So, four species categories were determined: very accidental species (Vac), with occurrence < 10 %, accidental species (Acc) with occurrences varying between 10-25 %, common species (Cmt) are found in 25-50 % of samples, and constant species (Cst) which detected in 50% or more of the total samples (Bigot and Bodot, 1973).
By the use of different nonparametric indices, which are calculated per field and per month, alpha diversity were evaluated: (i) number of species (S) that comprises the population present on a study site and at a given time, The estimation of species richness of this community was carried out with EstimateS software (Colwell, 2013), by calculationg the non-parametric asymptotic estimators (Chenchouni, 2017): (i) Jackknife 1 estimator (S Jack 1) : S Jack 1 = S + t 1 (v -1) / v), (ii) Chao 1 estimator (S Chao 1) : S Chao1 = S + [t 1 (t 1 -1)] / 2 (t 2 + 1)], with v: total number of samples, S: emperical species richness, t 1 : the frequency of species that found in only one sample, t 2 : species that found in only two samples.
The estimator values were given as mean ± SD (standard deviation).
The beta diversity of leaf beetle taxocenoses was compared between the cereal fields. Several qualitative (Jaccard and Sørensen) and quantitative (estimated Chao-Jaccard, Morisita-Horn and Bray-Curtis) indices were computed. Similarity values were obtained by using EstimateS software (Colwell, 2013).
Pearson correlation tests were applied between diversity parameters of leaf beetle communities for the three cereal fields and for the three months of the study by Past software. In addition, the spatiotemporal variation of diversity parameters (N, S and H') was tested by the multivariate statistic test GLM (generalized linear model) in R version 3.6.1 (R Core Team, 2016).
Variations in values of number of individuals and number of species between cereal fields and months are displayed in Figure 2.  (Table 1).

Diversity Parameters
Generally, barley was the richest cereal field with S = 4 and N = 390, followed by oats (S = 3, N = 569) and durum wheat (S = 2, N = 185). In contrast, the months of the study period had the same species richness value (S = 3).    The multivariate statistic test GLM indicated that the variation of diversity parameters was significant and different according to cereal fields (Table 3). In durum wheat, a significant decrease was observed for the individual number and the species richness (P < 0.001). In oats, the number of individuals increased significantly (P < 0.001), whereas, the H' value had a significant decrease (P < 0.001).
During months, the diversity parameters did not differ (P > 0.05).
For the entire study period, the relationship between diversity parameters showed only one significantly positive correlation between Simpson index (1 -D) and equitability (E) at P < 0.05. The same, except for Simpson index (1 -D), which showed a significant correlation with Shannon index (H') at P < 0.05, no significant correlation was observed with the other diversity parameters between cereal fields (P > 0.05) (Figure 3).

Species Richness Estimation
The non-parametric estimator (Chao 1) revealed that the estimated values of species richness for the different cereal fields were mostly close to the observed values of species richness, with a maximum difference of one species in the case of durum wheat (S obs = 2 ± 0.33, S Chao1 = 3.32 ± 0.34) and barley (S obs = 3, S Chao1 = 4 ± 0.37). Likewise, the non-parametric estimator (Jack 1) indicated that the estimated species richness values and the observed species richness values were barely different in durum wheat (S obs = 2 ± 0.33, S Jack1 = 3.32), barley (S obs = 3, S Jack1 = 4.64 ± 0.35) and oats (S obs = 3, S Jack1 = 4) ( Table 4).

Similarity Analysis
For all the quantitative and qualitative indices, similarity gave generally values > 39%, except for Bray-Curtis index (< 10%). The lowest similarities were observed between barley and oats, while the highest similarities marked durum wheat-barley and durum wheat-oats with values ranging from 75% to 99.7% (Table 5).

Phenology
Regarding the phenology of leaf beetle species in the cereal fields, O. melanopus was the species always appeared first, accompanied by O. gallaeciana in barley, and O. cyanella in oats. In addition, as far as the activity period is concerned, O. melanopus and O. gallaeciana had a longer activity period, from the 1 st week of April to late June in barley, whereas, in oats, the longest activity period was recorded by O. melanopus and O. cyanella (1 st week of April to late May). In durum wheat, both species O. melanopus and O. cyanella had almost the same activity pariod, which extended from the 1 st week of May to late June. The shortest activity period was registred for O. duftschmidi in barley (3 rd week of May). It is very important to note that, in durum wheat, the appearance of species was late, as compared to the other cereal fields (Figure 4).

DISCUSSION
The literature reveals few studies on the cereal leaf beetle (Oulema spp.) communities subservient to cereal field ecosystems. The present study targets to identify the leaf beetle fauna of the genus Oulema, and to determine the variation of the diversity parameters in space and time under semi-arid conditions in Eastern Algeria.
In general, the sampling of this insect group over three months in three different cereal fields ( (Popov et al., 2005). Likewise, both species O. melanopus and O. gallaeciana are the most represented in wheat and barley crops in the Northern part of Poland (Ulrich et al., 2004). In Turkey, the study carried out by Özdikmen (2011) showed the presence of three species of cereal leaf beetles: O. duftschmidi, O. gallaeciana and O. melanopus.
As far as the number of individuals and the number of species are concerned, the difference is important between the three cereal fields. The GLM test showed a high significant spatial variation (P < 0.001). The most abundant cereal fields was oats (569 individuals, 49.73%), whereas, the most speciose was barley (4 species). The lowest species richness and abundance were observed in durum wheat (N = 185, S = 2). Despite that the occurrence of each species varied from one cereal field to another, most species were classified constant. The species O.cyanella occurred commonly in durum wheat (Occ = 42.85%) and O. duftscmidi was very accidental (Occ = 7.69%) in barley. The study conducted by Rouag et al. (2012) provided the same species richness value in durum wheat. The difference in species could be attributed to their different trophic preferences; because barley and durum wheat fields were very close to each other. The floral choice of cereal leaf beetles was the subject of several studies. Price et al. (1980) indicated that barley and oats seem to be more attractive to cereal leaf beetles than wheat. In the same context, Philips et al. (2011) showed that the cereal leaf beetles feed on many species of grasses, and are considered as major enemies of oats, barley and common wheat (Triticum aestivum L.), which coroborrated to the findings of this study. However, differences in number of individuals could be explained by the practice of cereal monoculture and its intensity, it contributed significantly to the increase in the abundance of cereal leaf beetles (Kher et al., 2011). Furthermore, since they are phytophagous, cereal leaf beetles like leaves, where they lay their eggs and develop the different larval stages. In the case of our study, cultivators irrigated extensively the oats field; because it was intended to feed the animals, hence, the plant size increased remarkably, that is why oats field became a very favorable environment for their reproduction. A study conducted, in Romania, by Bucurean et al., (2012) reported that the intensity of the attack of O. melanopus on wheat was low, especially on the pubescent varieties, which are not preferable for the larvae, because they were so difficult to consume (mechanical resistance).
The species O. melanopus was the most abundant species in the three cereal fields, and the highest abundance value was recorded in oats with 362 individuals (63.62%). Dandurand (1976) underlined that the preferred host by O. melanopus was oats, which confirmed the results of the present study.
Based on Shannon index and Simpson index, there is an important spatiotemporal variation of the cereal leaf beetle community. In general, the values of diversity indices are low, ranging between 0.34 ± 0.09 and 0.94 ± 0.11. Barley represents the most diversified cereal field, and May is the most diversified month. Effectively, all species appeared in barley during May according to the phenogram. The low diversity can be attributed, from one side, to local harsh climatic conditions, because Tebessa and Batna belong to semi-arid bioclimatic stage, with a long dry period (April-October), according to Gaussen and Bagnouls diagram of the studied regions. On the other side, our study is interested only in cereals, there is a limitation of the prospected plants, the other plants are not taken into account, and otherwise, a large number of species belonging to the genus Oulema could be found. In this context Barbault (1981) reported that the increase in plant diversity leads to an increase in the diversity of phytophagous.
Beta diversity analyzed by quantitative and qualitative similarity indices expressed a high similarity between the three fields (> 50%) in most comparaison cases. This enormous similarity would be attributable mainly to the structure and the composition of cereal leaf beetle community subservient to the sampled cereal fields.
In the present study, two trapping techniques were used, which are the entomological net and the sight hunting. Based on the non-parametric estimators values (Chao 1 and Jackknife 1), it is very clear that the difference between the observed species richness and the estimated species richness is neglectible (one species at most), which signifies that the sampling effort was enough and missed species were almost inexistent.

CONCLUSION
In conclusion, this study provided important information on the cereal leaf beetles belonging to genus Oulema. This community has an important taxonomic richness, and spatiotemporal variations of alpha and beta diversity were recorded.