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Diversity and Characterization of Plant Parasitic Nematodes Associated with Cereals in Haiti

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Diversity and Characterization of Plant Parasitic Nematodes Associated with Cereals in Haiti Thesis Diversity and characterization of plant parasitic nematodes associated with cereals in Haiti Lesly Joseph Student number: 01700927 Promoter: Prof. Dr. Wim Bert A dissertation submitted to Ghent University in partial fulfilment of the requirements for the degree of International Master of Science in Agro- and Environmental Nematology Academic year: 2018 -2019 1 Diversity and characterization of plant parasitic nematodes associated with cereals in Haiti Lesly Joseph Nematology Research Unit, Department of Biology, Ghent University Ledeganckstraat 35, Ghent 9000, Belgium Abstract The diversity and the prevalence of plant parasitic nematodes associated with the cereals in Haiti have been assessed from samples collected in 35 agricultural fields: rice (18 samples), maize (11 samples) and sorghum (6 samples). Thirty (30) nematode species belonging to 12 genera have been identified. From the nematode species recovered, 20 nematode species from 12 genera were found associated with rice, 15 nematode species from 9 genera with maize and 9 nematodes species from 7 genera with sorghum. Five new species have been identified, from the following genera Helicotylenchus (2), Pralylenchus (1), Xiphinema (1) and Ditylenchus (1). From the nematode species recovered within the rice fields, Meloidogyne graminicola, Pratylenchus zeae, Helicotylenchus retusus were the most prevalent and abundant. Within the maize fields, Pratylenchus parazeae, Helicotylenchus erithrinae and Rotylenchulus reniformis were the most prevalent and abundant. In the sorghum fields, Pratylenchus parazeae, Helicotylenchus erithrinae and the criconematids were the most abundant nematode species but at low prevalence. This study confirmed the evidence that the prevalence and the density of plant parasitic nematodes are influenced by the crops. The sorghum cultivars in the investigated fields are poor host for the most damageable plant parasitic nematodes. Nine nematodes species (Tylenchorhynchus agri, Rotylenchulus reniformis, Pratylenchus parazeae, Pratylenchus zeae, Meloidogyne graminicola, Helicotylenchus erythrinae, Helicotylenchus retusus, Quinisulcius acutus, Mesocriconema sphaerocephalum) have been characterized morphologically and molecularly. This study provided the first molecular characterization and the first sequences for Helicotylenchus erythrinae, Helicotylenchus retusus and Quinisulcius acutus based on the 18S rDNA, D2-D3 and ITS1-5.8-ITS2 regions. The first sequences of the 18S rDNA gene for Tylenchorhynchus agri and Xiphinema basiri are provided from this research. This study presented an overview of the phylogeny of the sub-family of Telotylenchinae and suggested the combination of the genera Paratrophorus, Bitylenchus, Neodolichorhynchus, Sauertylenchus, Histotylenchus, Telotylenchus, Quinisulcius with Tylenchorhynchus to form a super genus of Tylenchorhynchus sensu lato. Key words: Plant parasitic nematode, cereals, phylogeny, species, Haiti, maize, rice, sorghum Introduction Plant-parasitic nematodes are omnipresent in all agricultural areas in the world. Globally, they are responsible of 10% of the total losses in the agricultural production chain (Savary et al., 2012; De Waele & Elsen, 2007). The monoculture and the conventional agriculture increase the density and the prevalence of plant of plant-parasitic nematodes (Coyne and al, 1999; Coyne and al, 2001). The biggest density of plant of plant parasitic nematodes has been observed in the ecosystem of the most cultivated crops in the 2 world (cereals, potatoes, soybean) (Sikora and al, 2018), which consequently induced more damages, more losses. According to FAO (2012), 30% of the total cereals produced in the world are lost. Taking into consideration the estimation of De Waele & Elsen (2007), plant parasitic nematodes are responsible of 3% of the total cereal lost worldwide. The cereals are staple foods for more than half of the human population, who are dependent at daily basis for their energy uptake (FAO, 2014). With the increasing human population, there is an increasing gap between the total cereals produced and the needs for those crops, which tend to affect the food security of the population of many third countries which are highly dependent on. In the Caribbean region, Haiti is among the countries with the highest rate of consumption of cereals, particularly rice (Sanou et al., 2013). Every year, on average an Haitian eat 40 kg of rice and on average 20 kg of maize and sorghum (Baro, 2002; Locher, 2001) . However, even though the cereals are crucial for the alimentation of the Haitian population, the country suffer a huge deficiency in the tonnage consumed compare to its production. In Haiti, more than 400 000 tons of rice are consumed yearly, while the annual production is around 172 000 tones, which represent 43% of the total needs and the rest are imported. The production of maize in Haiti is around 251 000 tons per year, which represent 90% of the total maize used for human consumption in the country (Dumazert & Ernest, 2017). Perhaps, the maize used as feeds are mainly imported, that constitute a barrier for the improvement of the livestock production in the country. Haiti is self-sufficient for sorghum supply for human consumption (ACF, 2017). However, with the increasing use of sorghum in beer production in Haiti, efficiency in the production should improve to prevent shortage. The agricultural system of Haiti is characterized by a low productivity and a low efficiency in the natural resources used in the production chain. Too many production factors are limited or missed in the production chain. The structural constraints intrinsic to this agricultural system, make plant parasitic nematodes not really regarded as a major problem that weaken the sustainability of this system of production. Therefore, plant parasitic nematodes don’t receive the attention they deserved in Haiti considering their known impacts in the agricultural sector worldwide. Furthermore, the lack of active nematologists and the low investment globally in agricultural researches from the government are also responsible of the sparsity of researches in the nematology field in Haiti. Consequently, most of the nematodes (Plant parasitic and free living) inhabitant the Haitian territory remain unknown to the scientific community. The first published report of plant of plant parasitic nematodes in Haiti was submitted by Crill (1973), exploring the diversity of plant parasitic nematodes associated with tomatoes in Haiti. Later, CIRAD (2005, 2012) presented Pratylenchus and Meloidogyne amongst the pests of yam and plantains in Haiti. Beside these two reports, no publication has been recovered about plant parasitic nematodes in Haiti. To our knowledge, none plant parasitic nematode from Haiti has already morphologically and molecularly characterized or link to molecular barcoding. Knowing that nematodes are omnipresent and caused important losses in the tropical regions where Haiti is, effective taxonomy strategy to identify the plant parasitic nematodes associated with the cash crops of Haiti at species level are an emergency now. The assessment of the diversity and the damage thresholds of the most abundant species will probably help to develop effective management strategy. The integrative taxonomy, combining traditional taxonomy and molecular barcode, is so far the best method to identify plant parasitic nematodes, as it combines the morphological and the genetic variations to determine the taxonomical status of a specimen or a population (Palomares-Rius et al., 2017). The molecular barcode in nematology is relevant, as it facilitates the identification of nematodes by non-experts in this field and globalized the research on the phylogeny of this order. The molecular barcode method could facilitate taxonomical work in the south countries like 3 Haiti, where expertise on nematode taxonomy are missing for the identification of the nematode morphologically. The general objectives of this study were to evaluate the density and the prevalence of the plant parasitic nematodes associated with rice, maize and sorghum and Provide morphological and molecular characterizations of the most important species recovered, including informative molecular barcodes for the partial 18S SSU rDNA, D2-D3 28S LSU rDNA and the ITS genes. Materials and methods Samplings and extraction During November 2018, samples of soils and roots where collected in 35 agricultural fields from maize (11 samples), rice (18 samples) and sorghum (6 samples) in Haiti (Fig 1). For each field, the geographical position coordinates were taken using a GPS Garmin. The samples were collected by digging out one plant and collect the soils attached and surrounding the roots system into a plastic bag. Soil samples were processed from a representative 200 ml using a modified Baermann funnel method. The nematodes recovered were fixed in DESS solution (DMSO-EDTA salt-saturated solution) in Haiti. The remaining part of each sample was brought to Belgium. In February 2019, 200 ml of soil from these samples were processed to extract the nematodes by using a modified Baermann funnel method for 72 hours and the nematodes recovered where fixed in Formalin. The Baunacke method was used for the extraction of the cyst nematodes in Belgium. Fig 1: Map presenting the locations of the samples collected in Haiti Density and prevalence value 4 The density of the different genera of PPN in each sample was calculated by pouring all the nematodes extracted into a counting dish
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