These sites are classified among the oldest rice-growing areas. Characterized by a humid climate with a unimodal rainfall, the agro-ecological zone of West Cameroon presents a very fertile agricultural land. Here, the data were collected in three localities: Dschang (05˚45'70.1''N - 10˚35'53''E, altitude: 1344 m) in the Menoua division, Ndop (06˚04'03.9''N - 010˚27'03.2''E, altitude: 1152 m) in the Ngo-Ketunjia division and Baïgom (04˚49'09.8''N - 011˚65'29.8''E, altitude: 1150 m) in the Noun division.

In this part of Cameroon, our research was carried out in three localities: Yaoundé-Nkolbisson (03˚51'57''N - 011˚27'11''E, altitude: 693 m) in the Mfoundi division, Ntui (04˚30'701''N - 011˚55'53''E., altitude: 509 m) in the Mbam-et-Kim division and Ebolowa (02˚56'52.1''N - 011˚07'9.7''E, altitude: 599 m) in the Mvilla division. In these localities, the climate is hot and humid, sub-tropical transition type, attenuated by altitude (Suchel, 1988). The annual temperature average is 23.5˚C contrasted between 16˚C and 31˚C depending on the season. Rainfall is 1650 mm per year [19]. The annual humidity average is 80% and varies during the day between 35% and 98%. Winds are wet and blow towards the southwest [19] [20].

In this region, we worked in two localities: Santchou (05˚15'47''N - 009˚58'10''E, altitude: 730 m) in the Menoua division and Tonga (04˚97'07''N - 010˚69'35''E, altitude: 1344 m) in the Ndé division. The climate prevailing in Santchou and Tonga is an equatorial type, characterized by the alternation of dry seasons and rainy seasons. With an average temperature of 23.6˚C contrasted between 19 and 31˚C depending on the season and 2478 mm of rain per year.

Direct planting was carried out on manually ploughed soil and each variety has been planted on 10 m². The spacing was 20 cm between the columns and 20 cm on the line. The distance between two varieties was 50 cm. Two weeks after planting, 300 kg∙ha⁻¹ NPK (20-10-10) were applied; 65 kg∙ha⁻¹ of urea spread at 46% of panicle initiation, that is to say 60 - 70 days after sowing according to the life cycle of the variety and 35 kg∙ha⁻¹ of urea at the flowering stage. Three manual weeding was carried out, respectively 15, 30 and 60 days after planting. The sampling step was two weeks.

Stem borers attack rates were calculated by reporting the number of white heads observed for each variety over the number of total panicles for each variety.

Attack   rate = ( Number   of   white   heads ) / ( Number   of   panicles )

The morphological identification, based on the dichotomous key [7] was first made. Forty seven insects, consisting of 46 larvae and 01 adult (obtained after raring) were collected from 04 localities in lowland and irrigated ecologies in Cameroon. These larvae and adult were preserved in absolute ethanol at −20˚C inside 2 mL Eppendorf tubes before genomic DNA extraction [21]. The genomic DNA extraction and SCAR-PCR analysis were carried out at the laboratory.

The extraction of the DNA was carried out by adding 600 μL of CTAB buffer (CTAB 2%, 0.1 M TRIS/pH 8, 0.02 M EDTA/pH 8, 1.4 M NaCl) in each tube containing the biological material then, each sample was incubated in a water bath at 60˚C for 30 minutes. Then, 600 μL of a solution containing a mixture in respective proportions 24/1 of chloroform and isoamylic alcohol was added to each tube and the resulting mixture was homogenized for 10 minutes. Centrifugation at 8000 rpm for 15 minutes was carried out, three phases were then distinguished: the aqueous phase was removed and transferred to a new 1.5 mL Eppendorf tube labelled as the old one. A volume of 450 μL of isopropanol gal was added to the volume of the aqueous phase taken. The contents of the tube were homogenized once more (a few seconds this time) and incubated at −20˚C for at least 30 minutes, for the precipitation of the nucleic acids. Centrifugation at 13,000 rpm for 20 minutes and at 4˚C followed, allowing the DNA to settle into a pellet at the bottom of the tube. The supernatant liquid was removed and the pellet was washed with 1 mL of 70˚ ethanol and then centrifuged at 13,000 rpm for 15 minutes and at 4˚C. Once the alcohol evacuated, the DNA pellet was dried and then suspended in 20 µL of sterile water. The extracted DNA was stored at −20˚C until use.

SCAR-PCR amplifies DNA sequences whose primers are designated from DNA sequenced fragments derived from RAPD (Random amplified polymorphic DNA) profiles.

Identification of the gall midge individuals was made by amplification of DNA sequences [14]. Each pair of primers used is specific to African Rice Gall Midge DNA (Table 1). The amplifications were carried out in a 25 μL reaction containing: the DNA extract, 2.5 μL, the 10 X TBE PCR buffer, 2.5 μL, 1 μL of the mixture of the deoxynucleotide triphosphates (dATP, dCTP, dGTP and dTTP), (10 mM), 1 μL of each primer of the SCAR primer pair considered concentrated at 10 Μm. 1 μL of MgCl₂ (25 mM), 0.1 μL of Taq polymerase (5 units/μL). The

amplifications were done in a thermocycler and the amplification program included: an initial denaturation step of the DNA at 94˚C for 4 min, 35 cycles each containing a denaturation step of the DNA at 94˚C for 1 min, a primer hybridization step at 60˚C for 1 min and an elongation step at 72˚C for 2 min, a final extension step at 72˚C for 7 min.

After PCR, the amplification products were separated by electrophoresis on a 2% agarose gel. The migration was made under the effect of an electrophoresis generator, at 100 volts for 1 hour, and after this, the gel is stained in a 0.5 mg∙mL⁻¹ ethidium bromide solution. For the interpretation of the results, a molecular weight marker was added during the migration.

During this study, Gall Midges were observed in the plots at the second week after planting. The frequency of these symptoms has been relatively low. These symptoms were observed only at the Yaoundé site (Figure 2(a)).

The first white heads were observed third week after planting on the Yaoundé site. Here, the most vulnerable varieties were: NERICA 3, NERICA 8, NERICA 9 and NERICA 13. The highest percentage of white heads (2.7%) was observed on the NERICA 3 variety, the lowest percentage (0.78%) was observed on the NERICA 9 variety (Figure 2(b)). Midge appeared at the vegetative stage of the host plant. At Ntui, the white heads did not appear until the ninth week after planting. FKR 60 had no white heads. In contrast, NERICA 3 had only one white head and NERICA 9 and 8 had the highest number of white heads (09 in total) (Figure 2(c)).

In the Ntui site, only Lepidoptera larvae were present. We observed that many Lepidopteron larvae can coexist in the same stem, while Dipterans larvae such as Diopsis sp. and Orseolia sp. were usually solitary (only one larvae per rice stem).

In Ndop site, no white head was observed. Rice farmers told us not to know about the white head phenomenon. However, some gall midges were observed in the plots where the rice was still at the vegetative stage.