Emilie Courvoisier

Yoann Bicaba

Xavier Colin
email: xavier.colin@ensam.eu

Multi-scale and multi-technical analysis of the thermal degradation of poly (ether imide)

Keywords:

The thermal degradation of PEI has been studied in wide ranges of temperature (between 180 and 250 °C) and oxygen partial pressure (between 0.21 and 50 bars). On one hand, the chemical ageing mechanisms have been analysed and elucidated by FTIR spectrophotometry and by differential calorimetry (DSC) on sufficiently thin PEI films (between 10 and 60 μm thickness) to be totally free of the effects of oxygen diffusion. As expected, and by analogy with other aromatic polymers of similar chemical structure, oxidation occurs preferentially on the methyl groups of the isopropylidene unit of the bisphenol A part, thus causing the disappearance of their characteristic IR absorption band at 2970 cm -1 and the growth of a new IR absorption band at 3350 cm -1 , attributed to alcohols. In addition, oxidation leads successively to a relative predominance of chain scissions and crosslinking, resulting in a non-monotonic change of T g . On the other hand, the consequences of oxidation on the elastic properties have been analysed and elucidated by micro-indentation on polished cross-sections of PEI plates of 3 mm thickness. The diffusion control of oxidation leads to the development of profiles of Young's modulus within the sample thickness, which correlate perfectly with the changes in chemical structure determined by FTIR spectrophotometry. However, the increase in Young's modulus in the superficial oxidized layer is not the direct consequence of oxidation but of a physical ageing.

Introduction

The competition in the aeronautical market depends both on economic and ecological issues such as the reduction of costs, on-board weight and fuel consumption. That is the reason why since its origin, the aerospace industry has been looking for new lighter materials with high thermomechanical properties. In this context, the use of composite materials made of organic matrix reinforced with glass or carbon fibers (OMC) has been amplified with the aim of progressively replacing the metallic materials. Initially designed for the fabrication of structural parts (wing, tail, fuselage, etc.), OMCs are now being considered for applications in increasingly harsh thermo-chemical environments, such as areas near heat sources (typically in environments of aircraft engine). However, they will be used only if their long-term durability is clearly demonstrated.

The use of thermoplastic matrices has several advantages, such as the integration of specific functions, but also the assembly of parts of the same or different nature. Among the thermoplastic matrices that can meet the aeronautical specifications, poly(ether imide) (PEI) appears as a serious candidate. Indeed, it is an amorphous polymer with a highly aromatic structure giving it not only high thermomechanical performances (T g ≈ 217 °C), but also a high chemical resistance to the most usual solvents (acetone, toluene, ethyl acetate, etc.) and aeronautical fluids (in particular, Skydrol). The thermal degradation of PEI has been little studied in the literature, almost exclusively in the rubbery state above 300 °C in inert or in slightly oxygenated atmosphere (typically in air).

The oxidation of PEI, but also of aromatic polymers with a similar chemical structure, occurs mainly at the methyl groups of the isopropylidene unit of the bisphenol A part, which contains the most labile H atoms [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Rivaton | Recent advances in bisphenol-A polycarbonate photodegradation[END_REF]. Oxidation leads to the formation of a wide variety of degradation products detected by FTIR spectrophotometry. These products include carbonyls, especially aromatic (at 1690 cm -1 ) and aliphatic ketones (1724 cm -1 ) and anhydrides (1840 and 1860 cm -1 ), as well as hydroxyls (3553 and 3514 cm -1 ) [START_REF] Rivaton | Recent advances in bisphenol-A polycarbonate photodegradation[END_REF][START_REF] Collin | Physicochemical and mechanical impacts of photo-ageing on bisphenol a polycarbonate[END_REF][START_REF] Rivaton | Comparison of the photochemical and thermal degradation of bisphenol-A polycarbonate and trimethylcyclohexane-polycarbonate[END_REF][START_REF] Kuroda | Degradation of aromatic polymers-II. The crosslinking during thermal and thermo-oxidative degradation of a polyimide[END_REF][START_REF] Corres | Thermal and thermo-oxidative degradation of poly(hydroxy ether of bisphenol-A) studied by TGA/FTIR and TGA/ MS[END_REF]. The disappearance of the IR absorption bands which are characteristic to the isopropylidene unit (C-CH 3 bond at 1186 cm -1 ) and methyl groups (C-H bond at 2870, 2933 and 2970 cm -1 ) has been also observed [START_REF] Collin | Physicochemical and mechanical impacts of photo-ageing on bisphenol a polycarbonate[END_REF][START_REF] Kuroda | Degradation of aromatic polymers-II. The crosslinking during thermal and thermo-oxidative degradation of a polyimide[END_REF].

Mechanisms of chain scission were also proposed to explain the formation of the wide variety of volatile products. Chain scissions occur at the C-C bonds of the isopropylidene unit, but also at the ether bonds between the phenyl and phthalimide rings [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Corres | Thermal and thermo-oxidative degradation of poly(hydroxy ether of bisphenol-A) studied by TGA/FTIR and TGA/ MS[END_REF][START_REF] Carroccio | Thermal degradation mechanisms of polyetherimide investigated by direct pyrolysis mass spectrometry[END_REF][START_REF] Lisa | Thermal and thermo-oxidative stability and probable degradation mechanism of some polyetherimides[END_REF][START_REF] Farong | The thermal stability of polyetherimide[END_REF]. They predominate largely over crosslinking at the beginning of exposure, but they become finally negligible in comparison to crosslinking at longer term [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Kuroda | Degradation of aromatic polymers-II. The crosslinking during thermal and thermo-oxidative degradation of a polyimide[END_REF][START_REF] Carroccio | Thermal degradation mechanisms of polyetherimide investigated by direct pyrolysis mass spectrometry[END_REF][START_REF] Dine-Hart | Oxidative degradation of a polyimide film: I. Initial studies[END_REF][START_REF] Augh | Degradation of continuous carbon fiber reinforced polyetherimide composites during induction heating[END_REF]. The bimolecular recombination of methylene radicals would be the main source of crosslinking.

The present article provides a detailed multi-scale and multi-technical analysis of the thermal degradation of PEI at lower temperatures and higher partial oxygen pressures than those studied until now in the literature. A peculiar attention is paid to the consequences of degradation on two key thermomechanical properties in the aeronautical field: the changes in the glass transition temperature and Young's modulus.

Experimental

The material under study is the PEI ULTEM ® 1010 supplied by SABIC company in the form of pellets. Plates of 3 mm thickness were molded by an injection machine from these pellets by the Pôle Européen de Plasturgie (PEP) located in Oyonnax (France). The plates were then cut with a LEICA RM2255 microtome in order to obtain sufficiently thin films (typically between 10 and 60 μm thickness) for ensuring a homogeneous oxidation throughout their thickness. The oxidation kinetics of PEI was studied in the rubber state at 220 and 250 °C, but also in the glassy state at 180 and 200 °C, under oxygen partial pressures ranged between 0.21 bar (in air-ventilated ovens regulated at ± 1 °C) and 50 bars (in autoclaves). All the films were periodically removed from the ageing chambers and cooled to room temperature in a desiccator containing silica gel for preventing any moisture recovery prior to being characterized. It is noteworthy that much thicker samples (plates of 3 mm thickness) were also exposed in air-ventilated ovens at 220 and 250 °C in order to put in evidence the development of oxidation profiles. In all cases, the consequences of oxidation were analysed by several complementary laboratory techniques from the molecular to the macroscopic scales, via the macromolecular and microscopic scales.

The changes in molecular structure were followed by FTIR spectrophotometry in a transmission mode. The FTIR spectra of the PEI films were measured before and after ageing for detecting and quantifying the disappearance of chemical groups initially present in the PEI matrix and the formation of the main oxidation products. The analyzes were carried out with a Perkin Elmer Frontier apparatus between 400 and 4000 cm -1 , after having averaged the 16 recordings obtained with a minimum resolution of 4 cm -1 . The thermal ageing of PEI leads to two major changes in the FTIR spectrum (Fig. 1). On the one hand, it can be seen the decrease in intensity of the IR absorption bands at 2970, 2935 and 2875 cm -1 , which are mainly characteristic of the C-H bonds of the -CH 3 groups of the isopropylidene unit in the early periods of exposure, but also of the C-H bonds of the -CH 2 -groups formed during thermal ageing at longer term (see Fig. 4). On the other hand, it is observed the appearance and growth of a wide IR absorption band centered at 3350 cm -1 and attributed to the elongation vibration of the O-H bond of alcohols.

Let us recall that the concentration C of these various products is related to the absorbance Abs of their IR absorption band according to the Beer-Lambert's law:

= × C Abs e ε ( 1
)
where e is the sample thickness (expressed in cm) and ε the coefficient of molar extinction whose averaged values are given in the literature:

ε(O-H of alcohols) = 90 L mol -1 . cm -1 [12,13] and ε(C-H of CH 3 ) = 42 L mol -1 . cm -1 [14].
Consequences of oxidation on the macromolecular architecture of PEI were followed by differential scanning calorimetry (DSC). The DSC thermograms of the PEI films were measured before and after thermal ageing for accessing the changes in the glass transition temperature (T g ). The analyzes were carried out with a TA Instruments Q1000 calorimeter between 25 and 300 °C with a heating rate of 10 °C.min -1 under a nitrogen flow. The value of T g was taken at the inflection point of the variation curve of the heat flux with the temperature.

Finally, the impact of thermal ageing on the elastic properties (in particular, on the Young's modulus) of PEI was determined by microindentation on the polished cross-sections of PEI plates of 3 mm thickness. The plates were cut in the thickness direction and embedded in a commercial acrylic KM-V resin, which was crosslinked for 12 h under primary vacuum at room temperature. Then, the plate crosssections were polished with a MECAPOL P320 device using silicon carbide abrasive papers of decreasing particle size (typically from 80 to 2400 granulometry). Finally, a mirror finish was obtained using diamond pastes of decreasing particle size (typically from 3 to 0.25 μm). The indentations were then performed with a CSM Instrument Micro Indenter equipped with a Vickers diamond tip of pyramidal geometry,

Table 1

Activation energy of the induction times (t i ) and the maximum rates (r ox ) of the disappearance of C-H bonds and the accumulation of alcohols in air between 200 and 250 °C.

Groups

E a of t i (kJ.mol -1 ) in air E a of r ox (kJ.mol with a force of 500 mN and a loading and unloading rate of 1000 μm min -1 . A pause of 5 s was systematically applied between the loading and unloading. The Indentation 4.37 operating software gives directly the value of the reduced modulus E r of the material, which is calculated according to Oliver & Pharr's method [START_REF] Collin | Physicochemical and mechanical impacts of photo-ageing on bisphenol a polycarbonate[END_REF][START_REF] Iqbal | Surface plasticization of poly(ether ether ketone)[END_REF][START_REF] Oliver | An improved technique for determining hardness and elastic modulus using load and displacement sensing indentation experiments[END_REF][START_REF] Clément | Détermination des propriétés mécaniques de céramiques poreuses par essais de microindentation instrumentée sphérique[END_REF]:

= E π S β A 2 r c ( 2
)
where S is the initial slope of the unloading curve, β is a shape factor depending on the indenter type (β = 1.012 for a Vickers tip) and A c is the contact area, between the indenter and the sample, projected perpendicularly to the indenter axis on the sample surface: A C = a 2 , a being the side length of the projected square. This latter quantity is also directly given by the operating software. It depends both on the penetration depth of the indenter and the indenter geometry.

Then, the local elastic modulus E was determined from the reduced modulus E r with the following equation:

= - - - E 1 ν E ν E 1 1 2 r i i 2 ( 3
)
where ν is the Poisson's coefficient of the virgin PEI: ν = 0.44 (supplier data), and ν i and E i are respectively the Poisson's coefficient and Young's modulus of the diamond indenter: ν i = 0.07 and E i = 1141 GPa. The global elastic modulus E S is the average value of the N local values E(j) constituting the micro-indentation profile:

∑ = = E N E j 1 ( ) S j N 1 (4)
In this study, E S was used as the Young's modulus of the sample after having checked, by uniaxial tensile testing, that it gives a very good estimate of this latter. Indeed, it has been found that their respective values differ only by a factor of 0.7.

Results and discussion

Changes in molecular structure

Fig. 2 reports the changes in the concentration of alcohols (at 3345 cm -1 ) and C-H bonds (2970 cm -1 ) during the thermal ageing of PEI in air between 200 and 250 °C. As expected, the temperature significantly accelerates the oxidation kinetics of PEI.

The consumption of C-H bonds is explained both by the release (in the form of methane) of CH 3 groups from the isopropylidene unit due to the thermolytic cleavage of the C-CH 3 bonds [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Corres | Thermal and thermo-oxidative degradation of poly(hydroxy ether of bisphenol-A) studied by TGA/FTIR and TGA/ MS[END_REF][START_REF] Carroccio | Thermal degradation mechanisms of polyetherimide investigated by direct pyrolysis mass spectrometry[END_REF][START_REF] Lisa | Thermal and thermo-oxidative stability and probable degradation mechanism of some polyetherimides[END_REF], but also by the preferential propagation of oxidation on the CH 3 groups. Indeed, let us recall that the dissociation energy of a C-H bond is much lower in a CH 3 group (≈ 414 kJ mol -1 ) than in an aromatic ring (≈ 465 kJ mol -1 ). In addition, the C-H bond is even more labile when the CH 3 group is located near a double bond or an aromatic ring. As an example, in the case of toluene, its dissociation energy is only 347 kJ mol -1 [START_REF] Calvert | Photochemistry[END_REF].

As an example, Fig. 3 compares the changes in the concentration of alcohols and C-H bonds in air at 220 °C. It is found that alcohols appear as soon as C-H bonds begin to disappear. More generally, the changes in these two species are perfectly correlated: the rates (of accumulation or disappearance) become maximum then slow down suddenly, to finally tend towards a quasi-constant value, after the same exposure durations. This result confirms that alcohols are formed by oxidation of the CH 3 groups as already described in the literature for polymers containing the bisphenol A unit in their chemical structure, in particular for PEI and polycarbonate (PC) [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Rivaton | Comparison of the photochemical and thermal degradation of bisphenol-A polycarbonate and trimethylcyclohexane-polycarbonate[END_REF].

In addition, the initial presence of an induction period on all the kinetic curves clearly indicates that the thermolysis of the isopropylidene unit is not the main initiation mechanism between 200 and 250 °C. Indeed, according to literature [START_REF] Colin | About a quasi-universal character of unstabilised polyethylene thermal oxidation kinetics[END_REF], we are rather in the case of a thermal oxidation mechanism initiated by its main propagation product: the hydroperoxide group POOH. Such a "closed loop" mechanism allows explaining why, during a more or less long induction period (depending both on the chemical structure of polymer and the agressiveness of ageing conditions), oxidation is not detectable by common spectrochemical techniques, but also why, at the end of this induction period, oxidation auto-accelerates sharply to finally reach its maximum rate. Let us recall that, at high temperature (typically above 200 °C), the decomposition of hydroperoxides is mainly unimolecular [START_REF] Colin | Determination of thermal oxidation rate constants by an inverse method. Application to polyethylene[END_REF].

In conclusion, these experimental results militate in favor of the thermal degradation mechanism proposed for PEI and PC at high temperature in the literature [START_REF] Carroccio | New vistas in polymer degradation. Thermal oxidation processes in poly(ether imide)[END_REF][START_REF] Rivaton | Comparison of the photochemical and thermal degradation of bisphenol-A polycarbonate and trimethylcyclohexane-polycarbonate[END_REF][START_REF] Carroccio | Thermal degradation mechanisms of polyetherimide investigated by direct pyrolysis mass spectrometry[END_REF][START_REF] Farong | The thermal stability of polyetherimide[END_REF], i.e. for an oxidation reaction initiated not only by the thermolytic breakdown of the C-C bonds of the isopropylidene unit, but mainly by the unimolecular decomposition of hydroperoxides, both resulting in the disappearance of CH 3 groups and the formation of alcohols. Fig. 4 summarizes these two degradation paths. It should be noticed that oxidation can also lead to the formation of ketones by rapid rearrangement (by β scission) of alkoxyl radicals (PO % ). Unfortunately, ketones are not detectable by FTIR spectrophotometry because of the initial presence of two broad IR absorption bands between 1680 and 1800 cm -1 , attributed to the elongation vibrations of the C-O bonds of imides.

Two ageing indicators were determined graphically for all the kinetic curves in order to clearly evidence the effects of temperature and oxygen partial pressure on the oxidation kinetics of PEI. They are: the induction time (t i ) marking the onset of the auto-acceleration stage, and the maximum oxidation rate (r ox ) corresponding to the maximum slope of the kinetic curves. First of all, these two indicators have been determined in air and plotted in the Arrhenius diagrams of Fig. 5.

Table 1 summarizes the activation energies of t i and r ox in air between 200 and 250 °C determined for C-H bonds and alcohols. As seen previously, the temperature significantly accelerates the oxidation kinetics of PEI. However, it reduces much more t i than it increases r ox . This large difference between the activation energies of both ageing indicators will be discussed later.

Then, to evidence the influence of the oxygen partial pressure on the oxidation kinetics of PEI, FTIR analyzes were performed on PEI films aged under oxygen partial pressures ranged between 0.21 (in air) and 50 bars, between 180 and 220 °C. As an example, Fig. 6 reports the changes in the concentrations of alcohols and C-H bonds at 220 °C. It is clear that the oxygen partial pressure accelerates the oxidation kinetics of PEI.

As an example, the values of t i and r ox determined for C-H bonds and alcohols at 200 °C were plotted as a function of the oxygen partial pressure in Fig. 7. It is noteworthy that these ageing indicators take comparable values for the two chemical species under consideration. In addition, the curves exhibit a hyperbolic shape and reach a plateau, which is a characteristic of the oxygen excess regime, under high oxygen partial pressures. In other words, t i and r ox become independent of the oxygen partial pressure above a critical value, denoted P C .

The values of P C , as well as the values of t i and r ox determined in oxygen excess between 180 and 220 °C are summarized in Table 2. It is noteworthy that the oxygen excess is not fully reached in air (i.e. under an oxygen pressure of 0.21 bar) at 200 and 220 °C. However, it appears clearly that P C is a decreasing function of temperature. Indeed, at 180 °C, oxygen excess is only reached beyond 20 bars.

Fig. 8 reports the Arrhenius graphs of t i and r ox in oxygen excess between 180 and 220 °C (see Table 2). The corresponding activation energies are given in Table 3. It can be seen that they are very similar for the two chemical species under consideration. As expected, for an oxidation reaction mainly initiated by the unimolecular decomposition of hydroperoxides, the activation energy of t i is higher than 140 kJ mol -1 , but not so much higher [START_REF] Zolotova | Mechanism of propagation and degenerate chain branching in the oxidation of polypropylene and polyethylene[END_REF]. In comparison, for a bimolecular decomposition, this value would have been appreciably lower: typically E a = 100-110 kJ mol -1 [START_REF] Khelidj | Oxidation of polyethylene under irradiation at low temperature and low dose rate. Part II-Low temperature thermal oxidation[END_REF][START_REF] François-Heude | Impact of oxygen transport properties on polypropylene thermal oxidation. Part 1: effect of oxygen solubility[END_REF].

In addition, the activation energy of r ox is located in the low range of the values usually reported in the literature for hydrocarbon polymers. Indeed, for a moderately oxidizable polymer such as PE, E a is quite high: E a ≈ 138 kJ mol -1 [START_REF] Khelidj | Oxidation of polyethylene under irradiation at low temperature and low dose rate. Part II-Low temperature thermal oxidation[END_REF]. In contrast, for highly oxidizable polymers such as PP or unsaturated elastomers, E a is significantly lower: E a ≈ 92 kJ mol -1 for PP [START_REF] François-Heude | Impact of oxygen transport properties on polypropylene thermal oxidation. Part 1: effect of oxygen solubility[END_REF] and E a ≈ 84 kJ mol -1 for HTPB [START_REF] Coquillat | Thermal oxidation of polybutadiene. Part I: effect of temperature, oxygen pressure and sample thickness on the thermal oxidation of hydroxyl-terminated polybutadiene[END_REF]. As feared, the presence of aromatic rings in the vicinity of the CH 3 groups greatly increases the oxidation sensitivity of these latter.

Finally, let us notice that the activation energies of t i and r ox take different values in oxygen default (for instance in air, see Table 1) due to the temperature dependence of the critical pressure P C .

Changes in macromolecular structure

The molecular mass is the structural variable commonly used for monitoring the changes in the macromolecular architecture of the polymer, i.e. chain scissions (denoted S) and crosslinking events (denoted X), during its chemical ageing. As each chain scission leads to a chain formation, whereas each crosslinking event leads to a chain disppearance, it can be written:

-= - S X M M 1 1 n n 0 ( 5
)
where M n0 and M n are the number average molecular masses before and after ageing respectively. However, in this study, it was impossible to measure directly the molecular mass of PEI after thermal ageing by a classical chromatographic (CES or GPC) or viscometric (in solution) method, because this polymer crosslinks and thus, becomes rapidly insoluble in its usual solvents, e.g. in dichloromethane [START_REF] Viallat | Viscosity behaviour of poly(ether imide) in solution: intramolecular interaction and intermolecular association effects[END_REF] and dimethylacetamide [START_REF]Polyimides and Other High Temperature Polymers: Synthesis[END_REF]. It was thus necessary to use an indirect method having already proved its worth for other aromatic polymers, for instance PEEK [START_REF] Richaud | Radiochemical ageing of poly(ether ether ketone)[END_REF]: differential scanning calorimetry (DSC). Indeed, DSC gives access to the value of M n if knowing the value of T g and applying the Fox-Flory's relationship [START_REF] Fox | Second-order transition temperatures and related properties of polystyrene. I. Influence of molecular weight[END_REF]:

⎜ ⎟ - =- ⎛ ⎝ - ⎞ ⎠ T T k M M 1 1 g g F F n n 0 0 (6)
where k FF is the Fox-Flory's constant (expressed in K.mol.kg -1 ), which

Table 4

Critical pressure (P c ), induction time (t i ) and maximum rate (r) for molar and macromolecular modifications in oxygen excess at 220 °C. is an increasing function of the chain stiffness. k FF was determined by three different methods: the Bicerano's relationship [START_REF] Bicerano | Prediction of Polymer Properties[END_REF], the free volume theory [START_REF] Richaud | Radiochemical ageing of poly(ether ether ketone)[END_REF][START_REF] Simha | On a general relation involving the glass temperature and coefficients of expansion of polymers[END_REF] and the copolymer theory [START_REF] Richaud | Radiochemical ageing of poly(ether ether ketone)[END_REF]. Its average value is: k FF = 295 ± 85 K kg mol -1 .

By combining Eq. ( 5) and Eq. ( 6), the following relationship was obtained between T g and the numbers of chain scissions and crosslinking events:

- = - T T k X S ( ) g g F F 0 (7)
It can be seen that T g decreases with the number of chain scissions, but increases with the number of crosslinking events. Fig. 9 reports the differences between T g and its initial value (T g -T g0 ) and between the numbers of crosslinking and chain scission events (X -S) for PEI films exposed in air between 200 and 250 °C. At 220 and 250 °C, two successive behaviors are clearly distinguished: first of all, an initial decrease in T g which reflects a relative predominance of chain scissions over crosslinking in the early periods of exposure; then, a strong increase in T g beyond its initial value, indicating that crosslinking becomes in turn predominant. These two phenomena (chain scissions and crosslinking) are highly thermo-activated. At 200 °C, it is suspected that this scenario is still valid, but it will only occur after a (long) induction period of about 5000 h. DSC analyzes were also carried out on PEI films after thermal ageing at 220 °C under different oxygen partial pressures ranged between 0.21 and 10 bars. The changes in (T g -T g0 ) and (X -S) are given in Fig. 10. It is found the same behavior as in air. However, the initial fall of T g is accentuated when increasing the oxygen partial pressure. In a general way, the oxygen partial pressure accelerates the kinetics of chain scissions and crosslinking up to a critical pressure of about 0.6 bar at 220 °C, a value which has been already determined, but with a better accuracy, by FTIR spectrophotometry. It is noteworthy that it was difficult to follow the PEI crosslinking above its initial value of T g , in particular under high oxygen partial pressure, due to the extreme brittleness of the PEI films.

The values of the induction time (t i ) of macromolecular modifications and the maximum rates of chain scission (r S ) and crosslinking (r X ) determined at 220 °C were plotted as a function of the oxygen partial pressure in Fig. 11. It can be seen that the values of r S and r X are comparable. In addition, the curves exhibit a hyperbolic shape and reach a plateau, which is a characteristic of the oxygen excess regime, above P C = 3 bars. The values of t i , r S and r X in oxygen excess at 220 °C are summarized in Table 4 where they are compared to the values obtained for the degradation products.

In Table 4, it can be seen that chain scissions are detected well before the disappearance of the CH 3 groups and the formation of alcohols, whatever the oxygen partial pressure. Thus, it can be deduced that they occur predominantly at the Ph-C(CH 3 ) 2 bonds in the initiation stages by thermolysis, but also by unimolecular decomposition of hydroperoxides (in particular, during the rearrangement by β scission of alkoxy radicals PO % ). Let us recall that this last reaction is in competition with the hydrogen abstraction leading to the formation of alcohols (Fig. 4). Concerning crosslinking, it becomes predominant over chain scissions after a longer exposure duration (800 h at 220 °C in air), i.e. when a sufficient quantity of alkyl P • and peroxyl PO 2

• radicals has been formed. It can thus be concluded that it results from the coupling of these radicals, as already proposed in the literature for PC [START_REF] Collin | Physicochemical and mechanical impacts of photo-ageing on bisphenol a polycarbonate[END_REF]. Examples of potential mechanisms of crosslinking for PEI are presented in Figs. 12-14. They all lead to a C-C bridge between two macromolecular chains. In the absence of oxygen, crosslinking could directly result from the coupling of primary P • radicals (Fig. 12). In oxygen default, as dialkyl peroxide bridges POOP cannot survive at the temperatures under study (typically for T ≥ 180 °C), they decompose almost instantaneously to form two PO % radicals which can rearrange in different ways and, in particular, into tertiary P • radicals by β scission (Fig. 13). These latter could then couple to form a crosslink node. The same reaction pathway could be proposed in oxygen excess, just after the rapid decomposition of the very unstable tetraoxide bridges POOOOP (Fig. 14).

Finally, it can be noticed that the macromolecular modifications are considerably slower than the disappearance of CH 3 groups or the formation of alcohols (see Table 4).

Consequences of oxidation on the elastic properties

The profiles of elastic modulus generated by oxidation in the thickness of PEI plates were determined by micro-indentation. As an example, the profiles obtained before and after 1966 h of exposure in air at 250 °C are reported in Fig. 15. As expected, the initial profile is almost flat. The corresponding average value E S of the elastic modulus is 5.0 ± 0.2 GPa. It is barely higher than the value of the Young's modulus determined by uniaxial tensile testing on dumbbell specimens: 3.5 ± 0.1 GPa. For this reason, in the present study, a correction factor of 0.7 has systematically been applied to the indentation modulus in order to access the value of the Young's modulus.

During thermal ageing, an increase in Young's modulus is observed in the superficial oxidized layer (Fig. 15). This increase correlates perfectly with the disappearance of CH 3 groups determined by FTIR spectrophotometry in an ATR mode (Fig. 16). In addition, the thicknesses of oxidized layer determined with these two techniques (i.e. micro-indentation and FTIR spectrophotometry) are comparable. They are of the same order of magnitude as the thicknesses of the brownish layer observed by optical microscopy on the free edges of the samples. As an example, Fig. 17 shows the micrographs obtained after four different times of exposure in air at 220 °C. It is clear that oxidation results in a browning of PEI.

The average values of the thickness of oxidized layer determined in air at 220 and 250 °C with these three techniques are summarized in Table However, the increase in Young's modulus is not a direct consequence of oxidation but, above all, of a physical ageing revealed by the appearance and growth of an endotherm of enthalpic relaxation (or structural relaxation) within the glass transition zone in the DSC thermograms (Fig. 18).

Thus, if PEI is initially in a glassy state (case of thermal ageings at 180 and 200 °C), the increase in Young's modulus is detected from the early periods of exposure. In contrast, if PEI is initially in a rubbery state (case of thermal ageings at 220 and 250 °C), the Young's modulus increase is detected only when T g becomes higher than the exposure • radicals) reduce also progressively the material ability to undergo a structural relaxation.

On this basis, a correlation was established between the local values of the Young's modulus, determined by micro-indentation before and after exposure of the PEI plates in air at 220 and 250 °C, and the corresponding values of the relaxation enthalpy determined by DSC (Fig. 20). It writes:

= + × E H 5. 2 0. 35 Δ (8)
The average value E OL of the Young's modulus of the oxidized layer, for PEI plates aged in air at 220 and 250 °C, was calculated with Eq. ( 4). These values were plotted as a function of the reciprocal T g in Fig. 21. It can be seen that all the points are placed around a single master straight-line whose the equation satisfies the relationship of Gilbert et al. [START_REF] Gilbert | Modulus-maps for amorphous polymers[END_REF]:

⎜ ⎟ = ⎛ ⎝ - ⎞ ⎠ E T E K α T T ( ) (0 ) 1 OL E g ( 9
)
where E(0 K) is the value of the Young's modulus at 0 K and α E is a constant. Thus, for PEI, it is found: E(0 K) = 4.1 GPa and α E = 0.18. It is noteworthy that this latter value is of the same order of magnitude as the value (α E = 0.3) proposed in the literature by Gilbert et al. [START_REF] Gilbert | Modulus-maps for amorphous polymers[END_REF], which would be, according to these authors, almost independent of the polymer structure.

Conclusion

The thermal degradation of the PEI matrix has been studied in the rubbery state (220 and 250 °C) and in the glassy state (180 and 200 °C) under oxygen partial pressures ranged between 0.21 and 50 bars, i.e. in domains of temperature and oxygen partial pressure still totally unexplored. The analysis at the molecular scale, by FTIR spectrophotometry, has allowed confirming the thermolysis and thermo-oxidation mechanisms proposed in the literature involving the CH 3 groups of the isopropylidene unit of the bisphenol A part, but also identifying alcohols as the main oxidation products. The analysis at the upper scales, by DSC then micro-indentation, has allowed showing, on the one hand, that chain scissions are majority at the beginning of exposure then crosslinking becomes predominant at longer term, whatever the oxygen partial pressure. On the other hand, the diffusion control of oxidation results in the formation of an oxidized superfical layer in which the elastic properties of PEI are significantly impacted. However, the increase in Young's modulus is not a direct consequence of oxidation, but it is due, above all, to a physical ageing.

In a next communication, a kinetic model will be derived from the oxidation mechanistic scheme established for PEI, and it will be interfaced with the different structure/property relationships (in particular, with Eq. ( 7) and Eq. ( 9)) identified in this study. This theoretical tool

Fig. 1 .

 1 Fig. 1. Changes in the regions of C-H (a) and O-H bonds (b) in the FTIR spectrum of PEI during its thermal ageing in air at 250 °C.

Fig. 2 .Fig. 3 .

 23 Fig. 2. Changes in the concentration of alcohols (a) and C-H bonds (b) during the thermal ageing of PEI in air at 200 (in pink), 220 (violet) and 250 °C (green). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4 .

 4 Fig. 4. Thermal degradation mechanism of PEI initiated by two competitive paths [1,4,7,9]: breakdown of C-C bonds of the isopropylidene unit (left) and unimolecular decomposition of hydroperoxides (right).

Fig. 5 .

 5 Fig. 5. Arrhenius diagram of the induction times (a) and the maximum rates (b) of the disappearance of C-H bonds (■) and the accumulation of alcohols (•) in air between 200 and 250 °C.

Fig. 6 .Fig. 7 . 2 3 Fig. 8 .

 67238 Fig. 6. Changes in the concentration of alcohols (a) and C-H bonds (b) during the thermal ageing of PEI under an oxygen pressure of 0.21 bar (in violet), 0.4 bar (black), 1 bar (red), 3 bars (orange) and 5 bars (green) at 220 °C. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 9 .

 9 Fig. 9. Differences between the glass transition temperature and its initial value (T g -T g0 ) (a) and between the numbers of crosslinking and chain scission events (X-S) (b) during the thermal ageing of PEI in air at 200 (in pink), 220 (violet) and 250 °C (green). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 10 .Fig. 11 .

 1011 Fig. 10. Differences between the glass transition temperature and its initial value (T g -T g0 ) (a) and between the numbers of crosslinking and chain scission events (X-S) (b) during the thermal ageing of PEI under an oxygen pressure of 0.21 bar (in violet), 0.4 bar (black), 3 bars (red) and 10 bars (blue) at 220 °C. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

4 Fig. 12 .

 412 Fig. 12. Termination by coupling of P % radicals leading to the formation of a C-C bridge between two macromolecular chains and thus, to a crosslinking node.

Fig. 13 .

 13 Fig. 13. Termination by coupling of P % et PO 2 % radicals leading to the formation of a C-C bridge between two macromolecular chains and thus, to a crosslinking node.

Fig. 14 .

 14 Fig. 14. Termination by coupling of PO 2 % radicals leading to the formation of a C-C bridge between two macromolecular chains and thus, to a crosslinking node.

Fig. 15 .

 15 Fig. 15. Profile of the Young's modulus in the thickness of the PEI plate before (in black) and after (red) 1966 h of exposure in air at 250 °C. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 16 .

 16 Fig. 16. Comparison of the degradation profiles determined throughout the thickness of a PEI plate by FTIR spectrophotometry (•) and micro-indentation (▲) after 1966 h of exposure in air at 250 °C.

Fig. 17 .

 17 Fig. 17. Examination by optical microscopy (with a magnification of × 2,5) of the superficial oxidized layer of PEI after different durations of exposure in air at 220 °C.

Fig. 18 .

 18 Fig. 18. DSC thermogrammes of PEI films after 535 h and 2000 h of exposure in air at 250 °C.

Fig. 19 .Fig. 20 .

 1920 Fig. 19. Changes in the enthalpy (a) and the temperature of the endothermic peak (b) during the thermal ageing of PEI in air at 180 (black), 200 (pink), 220 (violet) and 250 °C (green). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 21 .

 21 Fig. 21. Changes in the Young's modulus of the oxidized layer of PEI plates in fonction of its reciprocal T g before (•) and after exposure in air at 220 (■) and 250 °C (▲).

Table 3

 3 Activation energy of the induction times (t i ) and the maximum rates (r ox ) of the disappearance of C-H bonds and the formation of alcohols in oxygen excess between 180 and 220 °C.

	Groups	E a of t i (kJ.mol -1 ) in O 2	E a of r ox (kJ.mol -1 ) in O 2
		excess	excess
	C-H bonds	153	97
	O-H bonds (alcohols)	171	89

Table 5

 5 Average thickness of oxidized layer of PEI determined by micro-indentation, FTIR spectrophotometry or optical microscopy after different durations of exposure in air at 220 and 250 °C.

	T (°C)	t (h)	l ox (μm)
	220	429	275 ± 25
		667	400 ± 50
		1270	750 ± 50
		1985	750 ± 50
	250	94	225 ± 25
		670	350 ± 75
		1273	490 ± 60
		1966	550 ± 50

will allow us predicting not only the degradation state of PEI, but also the changes in its thermomechanical properties during the thermal ageing. The validity of this tool will be checked from all the experimental data collected in this study.


Anne Griffond-Boitier

Armelle Couillet

Thibaut Desmettre

Marc Puyraveau

Marc Puyraveau L'

téléphonique : l'apport du géographe

come


Larson Boundenga

Illich Cyr Moussadji

Manfred Mombo

Barthélémy Ngoubangoye

Jean Bernard Lekana-Douki

Jean-Pierre Hugot

Illich Manfred Mombo

Diversity and prevalence of gastrointestinal parasites in two wild Galago species in Gabon

Keywords:

à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Pathogens are playing a central role in ecosystems and community biodiversity due to their impact on the dynamic of populations growth and regulation [START_REF] Hochachka | Density-dependent decline of host abundance resulting from a new infectious disease[END_REF][START_REF] Hudson | The Ecology of Wildlife Diseases[END_REF]. A review of the literature shows that about 90% of the parasites recorded in Man include another mammal species in their cycle [START_REF] Ashford | Parasites of Homo sapiens: An Annotated Checklist of the Protozoa, Helminths and Arthropods for Which We Are Home[END_REF][START_REF] Hugot | Biodiversity in helminths and nematodes as a field of study: an overview[END_REF]. If animal's infectious diseases and parasitosis are a major threat for the conservation of wild species they also are a potential threat for human health [START_REF] Lyles | Infectious disease and intensive management: population dynamics, threatened hosts, and their parasites[END_REF] and the increasing conquest of the rain forest environments by men heighten the probability of parasite host switching. Phylogenetic proximities between Man and other hosts like NHPs (non-human primate species) are an important factor in pathogens exchanges [START_REF] Wolfe | Origins of major human infectious diseases[END_REF]. Consequently, it is important to determine the diversity of pathogens circulating in wildlife, and particularly among species endangered because of human activities [START_REF] Daszak | Emerging infectious diseases of wildlifethreats to biodiversity and human health[END_REF]. In this context, the data on patterns of parasitic infections in wild primate populations may provide an estimation of population health and allow to assess and manage disease risks [START_REF] Gillespie | Noninvasive assessment of gastrointestinal parasite infections in free-ranging primates[END_REF].

Gabon is a central African country whose forests covers 85 of the area and are among the richest in Africa in terms of animal diversity. The estimated fauna comprises > 190 mammal species, including at least 20 species of monkeys, > 600 bird species, 86 reptile species and 100 species of amphibians [START_REF] Alonso | Gamba, Gabon: biodiversity of an equatorial African rainforest[END_REF][START_REF] Pauwels | Reptiles du Complexe d'Aires Protégées de Gamba, sud-ouest du Gabon[END_REF]. Several species of nocturnal NHPs are living in Gabon including: Galago demidoff G. Fischer, 1806 andG. thomasi Elliot, 1907 [START_REF] Wilson | Mammal Species of the World: A Taxonomic and Geographic Reference[END_REF]. Galagos (bushbabies) are threatened by an increasing fragmentation of their habitat due to human activities, but last studies on these species were done long time ago and very few data are available about their parasites. In the following, using faecal samples collected in these hosts in Gabon, we examined the diversity and prevalence of gastrointestinal parasites from these two closely related species, in different study sites.

Materials and methods

Ethics statement

The study was conducted in Gabon. All samples were collected in ⁎ Corresponding author at: Groupe Évolution et Transmission Inter-espèces de Parasites (GETIP), Unité Évolution, Épidémiologie et Résistances Parasitaires, CIRMF, BP769 Franceville, Gabon.

⁎⁎ Corresponding author.

E-mail addresses: boundenga@gmail.com (L. Boundenga), hugot@mnhn.fr (J.-P. Hugot).

T areas excluding national parks and protected areas and with the authorization of the Gabonese Ministries of Water and Forestry, Higher Education, Scientific Research and Innovation (N°AR0031/09/MENE-SRESI/CENAREST/CG/CST/CSAR). During faecal collections, animal welfare was taken into consideration and each animal was released into its environment after the collect of biological material.

Sites for collection of NHPs

Research of gastro-intestinal parasites was conducted from February 2016 to January 2017 (11 months) on two different host species and two sites located in the province of Haut-Ogooué, Southeast Gabon (Fig. 1): the forest of the Primate Centre (CDP) of the "Centre International de Recherches Médicales de Franceville" (CIRMF) and the "Djoumou forest" (DJM) located between 30 and 40 km from Franceville. In CDP, forest habitats are divided into various relictual areas submitted to disturbances resulting from human activities. DJM forest covers a wider area than CDP and human presence is limited to traditional plantations and hunter's paths. Galagos were collected using Tomahawk traps [START_REF] Duplantier | Les rongeurs myomorphes forestiers du nord-est du Gabon: structure du peuplement, démographie, domaines vitaux[END_REF][START_REF] Malcolm | Comparative abundances of Neotropical small mammals by trap height[END_REF] placed in trees for six consecutive nights at different heights. All animals collected were kept in black room (without light) until faecal samples may be collected. Galagos species identification was performed using [START_REF] Kingdon | Kingdon Field Guide to African Mammals[END_REF] and [START_REF] Wilson | Mammal Species of the World: A Taxonomic and Geographic Reference[END_REF]. Parameters such as host species, sex, age appraisal and sampling season were documented during the collections.

Faecal samples collection and examination

A total of 147 bushbabies: G. demidoff (n = 83) and G. thomasi (n = 64) were examined. Faecal samples were analyzed via sodium nitrate flotation and faecal sedimentation following the protocol of [START_REF] Gillespie | Noninvasive assessment of gastrointestinal parasite infections in free-ranging primates[END_REF]. Detection and counting of parasite eggs, larvae, and cysts was performed using a compound microscope (objective ×10). We used the ×40 and ×100 objectives to take pictures (Fig. 2). Identification of parasites is based on eggs shell and nucleus morphology according to [START_REF] Chitwood | An Introduction to Nematology: Section 1: Anatomy[END_REF], [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF], [START_REF] Ashford | Parasites of Homo sapiens: An Annotated Checklist of the Protozoa, Helminths and Arthropods for Which We Are Home[END_REF]. Together with [START_REF] Combes | Interactions Durables. Ecologie et évolution du parasitisme[END_REF] and [START_REF] Anderson | Nematode Parasites of Vertebrates[END_REF], the same books were used to obtain information about the parasites biology and their host spectrum.

Statistical analyses

Statistical analyses performed using R software. Infestation rates between species, sexes and age groups calculated and compared using the Chi-square (χ 2 ) method. Differences were considered statistically significant at 0.05 confidence level.

Results

Here after we consider successively the different parasite taxa in relation to the importance and variations of their prevalence in different hosts (Fig. 2 and Tables). We also try to relate these variations to what we know about the cycle, host specificity and taxonomic status of the parasites. The identification of the parasites collected during our study is based on the light microscope observation of the eggs or cysts found in the faecal samples and by comparison with similar documents found in the literature. This method doesn't allow a precise determination of the parasite at the species level and in the following we identify the positive samples at the generic level only. The genus name used is that of the parasite of the same type (order, suborder, family) also found in other Primates, including Man. When several names were available we chose the most probable of them in relation to the particularities of the parasite cycle and the systematic position of the hosts. For most of the parasites, we also had to consider the possibility that each of the taxonomic units distinguished in this study may include different parasite species; for instance following which locality and/or host species is considered. This is the reason why we are systematically using here after the abbreviation "spp".

Ancylostoma spp.

The prevalence values are comprised between 7.5 and 10 and are twice as high in CDP than in DJM for both host species (Table 1). The prevalence is higher in females than in males (5.1 vs 13.2) and more than ten times higher in the young than in the adults (22.2 vs 2.1) (Table 3). After [START_REF] Anderson | Nematode Parasites of Vertebrates[END_REF] infection with hookworms is due to the penetration of larvae (L3) through the skin or by direct ingestion of dirt or fresh vegetables containing filariform larvae. They are parasite of the small intestine of mammals. Such parasites where identified as Necator spp. in both gorillas and chimpanzees by [START_REF] Graber | Parasites internes des primates de la République Démocratique du Congo (d'après la collection Cassard-Chambrom 1956-1960). Rôle pathogène-diagnostic prophylaxie[END_REF] in Congo. [START_REF] Kouassi | Diversity and prevalence of gastrointestinal parasites in seven non-human primates of the Taï National Park, Côte d'Ivoire[END_REF] also signal hookworms, identified as Necator sp. or Ancylostoma sp., in several different guenon species in the Tai forest (Ivory Coast) sometimes with a high prevalence value (> 70). Because [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF] signal Ancylostoma spp. in G. demidoff and G. gabonensis in the Democratic Republic of Congo with a high prevalence: 25.3 and 50 respectively; also because [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF] remarks that Ancylostoma eggs contain 4 blastomers only when laid, whereas Necator eggs contain 8, the parasites observed in our work must more likely be identified as Ancylostoma spp.

Ascaris spp.

This parasite exhibits one of the highest prevalence values observed in our study: 63.2 in G. thomasi and in CDP. For both host species the prevalence is higher in CDP than in DJM (Table 2) and in the males than in the females (Table 3). The Ascaridoidea are parasitic for vertebrate hosts: contamination results from ingesting food (vegetables or fruits) soiled in contact with earth or by drinking water. Genus Ascaris spp. are the single taxa in this group recorded in Primate hosts and, within the NHPs, they were observed in apes (gorillas and chimpanzees) [START_REF] Graber | Parasites internes des primates de la République Démocratique du Congo (d'après la collection Cassard-Chambrom 1956-1960). Rôle pathogène-diagnostic prophylaxie[END_REF] and in G. demidoff by [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF] with a prevalence of 37.04. In both publications the authors identify the parasites collected in Primates as A. lumbricoides, the parasite of Man. However this identification looks dubious because A. lumbricoides is generally described as highly specific and unable to maintain itself in another host than man [START_REF] Anderson | Nematode Parasites of Vertebrates[END_REF][START_REF] Ashford | Parasites of Homo sapiens: An Annotated Checklist of the Protozoa, Helminths and Arthropods for Which We Are Home[END_REF].

Balantidium spp.

For both host species the prevalence for this parasite is not very different (19.3 vs 15.6) (Table 1). The prevalence is markedly highest in CDP than DJM, relatively close between males and females (15.2 vs 20.6), and very different between adults or young (24.7 vs 4.0) (Tables 2 and3). Balantidium coli is a cosmopolitan parasite of the large intestine of the pig, but it is also found in vicarious hosts: Primates (macaques, guenons), Rodents (especially rats), dogs, cats and cattle. The cysts resist > 3 months in the wild and can be carried by flies [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF]. This parasite has been identified in several species of African guenons [START_REF] Kouassi | Diversity and prevalence of gastrointestinal parasites in seven non-human primates of the Taï National Park, Côte d'Ivoire[END_REF] living in the rain forest, in which it appears to be relatively frequent and abundant. These characteristics, the indisputable ubiquity of this parasite and its proven presence in the same habitats make its presence in the galagos very likely.

Dicrocoelium spp.

For both host species the prevalence values are similar and relatively weak (< 10) (Table 1); the contrast between the two sites is marked: prevalence is two to eight times higher in CDP (Table 2); the males are twice as parasitized as the females; the prevalence values are identical in adults or young: around 8 (Table 3). Dicrocoelium spp. live mainly in the bile ducts and gall bladders of domestic and wild ruminants but occasionally parasitize primates including Man. They use insects, especially ants, as intermediate hosts [START_REF] Kouassi | Diversity and prevalence of gastrointestinal parasites in seven non-human primates of the Taï National Park, Côte d'Ivoire[END_REF]. Ingesting infected ants or food contaminated by ants infects monkeys. This way of contamination is accessible to galagos, which are insectivorous. [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF] describes the true parasitism by Dicrocoelium as exceptional in Man, but signals the frequent finding of eggs in transit, in the feces. This remark may very probably be extended to other primates and is corresponding with our present observation.

Table 1

Prevalence of eleven parasite taxa collected in Galago demidoff or G. thomasi during the period of study. Prevalence comparison between host species, computed using the chi-square test. (N = number of positive specimens; P = prevalence; S = +. test significant; Gd + higher prevalence in G. demidoff; Gt + higher prevalence in G. thomasi).

Parasites species

Galagoides 3.5. Entamoeba spp.

For both host species the prevalence values are similar and relatively weak (< 10) (Table 1); the prevalence is very similar in CDP or DJM and higher in females or young (Tables 2 and3). Following [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF] about ten different species of Entamoeba have been identified in Man. A part of them are apparently non pathogenetic. Infection occurs orally, from drinking water, food, etc. E. histolytica is the most frequent, predominantly infecting humans and other primates and distributed worldwide. E. coli is identified as a parasite of chimpanzees and baboons by [START_REF] Howells | Patterns of gastro-intestinal parasites and commensals as an index of population and ecosystem health: the case of sympatric western chimpanzees (Pan troglodytes verus) and Guinea baboons (Papio hamadryas papio) at Fongoli, Senegal[END_REF]; the same species was recorded in Eulemur fulvus E. Geoffroy, 1796 from Madagascar, by [START_REF] Clough | Host intrinsic determinants and potential consequences of parasite infection in free-ranging red-fronted lemurs (Eulemur fulvus rufus)[END_REF]. If according to the above, finding Entamoeba spp. in galagos is not surprising more investigations are needed to precise which species is/are present in these particular hosts.

Gongylonema spp.

This parasite is present in both host species with a relatively low prevalence: 4.7 and 6 in G. thomasi and G. demidoff respectively (Table 1). It is absent from CDP and more frequent in females or young (Table 3). Members of the genus occur embedded in the mucosa of the anterior region of the gut of birds and mammals [START_REF] Anderson | Nematode Parasites of Vertebrates[END_REF]. The eggs are evacuated in the outside environment. If ingested by coprophagous insects (cockroaches, dung beetles, etc.) the larvae continue their development in the haemocoel of insects or become encapsulated in their muscles [START_REF] Quentin | Gongylonema congolense Fain, 1955, (Nematoda, Spirurida) synonymie et biologie larvaire, réactions cellulaires de l'insecte hôte intermediare[END_REF]. Definitive hosts are contaminated by accidentally swallowing these insects [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF]. G. pulchrum is the most frequent and cosmopolitan species. This nematode is usually a parasite of the digestive tract of cattle and swine but Man infestations have been reported in most parts of the world. The high frequency of this parasite in tropical areas, its ubiquity including Man and the fact that insects are intermediary hosts makes plausible its presence in galagos. More investigations are needed to precise which species is/are present in these particular hosts.

Lemuricola spp.

The samples collected in G. thomasi are negative all of them. Three individuals of G. demidoff only, out of 83 animals examined, were found positive and all of them were captured in DJM. An Oxyurid identified as Enterobius sp. is recorded by [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF] in a free-living galago: G. demidoff in Congo with a prevalence of 11.1. In a previous work [START_REF] Hugot | Morphology and Morphometrics of three oxyurids parasitic in primates. Description of Lemuricola microcebi n. sp[END_REF] oxyurids were found and described in a closely related host: G. senegalensis E. Geofffroy. But this animal was dead in captivity in the Faculty of Medicine of Paris where it was living in close proximity to a colony of Microcebus murinus (Miller) in a breeding lab. The parasites collected from the galago were morphologically very similar to Lemuricola microcebi,Hugot et al., 1995 considered to be specific for M. murinus. Finally because of the low numbers of parasites available for examination, it was decided impossible to choose between: either a transfer from the microcebes to the galago or the possibility that the two samples actually belong to two different species. Until more material or information could be obtained, it was proposed to refer to the parasites of the galago as Lemuricola sp. The results exposed above are giving some consistence to the existence of one or several particular and new parasite species in the galagos.

Table 2

Prevalence of parasite taxa collected in Galago demidoff or G. thomasi during the period of study following the site of collection. Comparison of prevalence between the two sites computed using the chi-square test. (N = number of positive specimens; P = prevalence; S = +. test significant; CDP = S.1. Primatology Center forest; DJM = S.2. Djoumou forest; + higher prevalence in CDP or DJM). With a prevalence of 39.1 for G. thomasi this parasite reach the highest value after Ascaris spp. for this host and one of the highest values observed for a parasite in DJM: 19 (Tables 1 and2). For both host species the prevalence is higher in CDP than in DJM (Table 2) and in the young than in the adults (Table 3). After [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF]: several Oesophagostomum spp. are present in African Primates like macaques, baboons, guenons and gorillas; all of them may contaminate Man who is considered a substitution or abnormal host; the oesophagostomiasis are more common in forest areas. Oesophagostomiasis has no vector, percutaneous infection has never been reported and it is believed that contamination occurs by oral-faecal ingestion of L3 larvae, both for Man or animals. Whether or not parasite transmission is specifically waterborne, foodborne, or both is not elucidated [START_REF] Ziem | Controlling Human Oesophagostomiasis in Northern Ghana[END_REF]. Future investigations will have to determinate if the parasite(s) observed in this study result of contamination by monkey or ape parasite(s) or are host specific for the galagos.

Strongyloides spp.

The prevalence of this parasite is more than five times higher in G. demidoff (62.7) than in G. thomasi (10.9) (Table 1) and for both hosts species the prevalence is higher in CDP than in DJM (Table 2). The prevalence is higher in males than in female, and in adults than in young (Table 3). Non-human primates are known to be the major hosts of Strongyloides spp. [START_REF] Egido | The prevalence of enteropathy due to strongyloidiasis in Puerto Maldonado (Peruvian Amazon)[END_REF][START_REF] Kouassi | Diversity and prevalence of gastrointestinal parasites in seven non-human primates of the Taï National Park, Côte d'Ivoire[END_REF] and particularly for Strongyloides fulleborni Linstow, 1905, a parasite of the monkeys of Asia and Africa. [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF] recorded a parasite identified as S. stercoralis [START_REF] Bavay | Sur tanguillule stercorale[END_REF] in G. demidoff in Congo with a prevalence of 14.81. These nematodes are found both in populations living in forested and urban areas. Some "strains" seem perfectly adapted to Man and can therefore be maintained outside the presence of monkeys. It should be noted that in these cases of human infestation, eggs and not larvae are found in stools. Future investigations will have to determinate if the parasite(s) observed in this study result of contamination by monkey or ape parasite(s) or are host specific for the galagos.

Trichostrongylus spp.

The prevalence for this parasite is much higher in G. thomasi (23.4 vs 1.2) and it is observed in DJM, only. The prevalence is about three times higher in the females than in the males (16.2 vs 6.3), and in the young than the adults (22 vs 5.2) (Table 3). According [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF], Trichostrongylus spp. are cosmopolitan and above all parasites of Ruminants, Lagomorphs and Rodents sometimes also from Equidae. In Africa these parasitosis are known in Man and monkeys: the eggs are eliminated in the stools and the faeco-oral contamination of Primates considered resulting from absorption of plants soiled by droppings of domestic animals [START_REF] Munene | Helminth and protozoan gastrointestinal tract parasites in captive and wildtrapped African non-human primates[END_REF]. This modality of infection looks unlikely or almost difficult for galagos, which are strictly arboreal. However Trichostrongylus spp. were recorded both in G. demidoff and G. gabonensis in Congo by [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF] with a prevalence of 50 for each host species. The fact that the prevalence is greater on the site where interactions with Man are less frequent (DJM) doesn't make very plausible a contamination by human parasites. A precise identification of the parasite(s) identified as Trichostrongylus spp. is necessary before we can interpret this observation.

Table 3 Distribution and prevalence of parasite species reported to the age or gender of galagos collected during the period of study. (N = number of positive specimens; P = prevalence; S = +, test significant; + higher prevalence in males (M), females (F), adults (A) or young (Y). The prevalence for this parasite is one of the lowest in both host species: 3.6 vs 9.4 (Table 1). Its prevalence is higher in CDP and quite identical in males, females, and adults or young (≈6) (Tables 2 and3). The eggs of this parasite localized in the small intestine are evacuated in the external environment with the stool. The eggs are extremely resistant to atmospheric and chemical agents and must stay in the ground during 6 to 12 months before to be infective. Man may be infested with fruits or vegetables growing on ground and raw [START_REF] Golvan | Atlas de Parasitologie (Schémas explicatifs d'épidémiologie)[END_REF]. About 60-70 species of whipworms have been recorded from mammals (primates, artiodactyls, rodents, lagomorphs, shrew, felids and foxes) and they are cosmopolitan [START_REF] Anderson | Nematode Parasites of Vertebrates[END_REF]. T. trichiura is a parasite of humans and other primates and is regarded as the second most common parasitic infection in humans in the tropics. [START_REF] Ooi | Comparative study of Trichuris trichiura from non-human primates and from man and their difference with T. suis[END_REF], described specimens from Macaca fuscata and Papio papio as T. trichiura. Future investigations will have to determinate if the parasite(s) observed in this study result of contamination by Man, monkey or ape parasite(s) or are host specific for the galagos.

Discussion

Recently several works were published regarding the parasitic fauna of African NHPs [START_REF] Landsoud-Soukate | Intestinal parasites of sympatric gorillas and chimpanzees in the Lopé reserve. Gabon[END_REF][START_REF] Clough | Host intrinsic determinants and potential consequences of parasite infection in free-ranging red-fronted lemurs (Eulemur fulvus rufus)[END_REF][START_REF] Nath | Prevalence of parasitic infection in captive non human primates of Assam state zoo, India[END_REF][START_REF] Kalousová | Gastrointestinal parasites of savanna chimpanzees (Pan troglodytes schweinfurthii) in Ugalla, Tanzania[END_REF] but only one of them concerned the galagos: [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF]. Unfortunately, if this work is providing a first sight on the diversity of the gastro-intestinal parasite fauna in two galago species, the observations are founded on a relatively low number of specimens (N = 30) captured in not least than seven different stations of captures. Logically considering this, the positive results are not identified following their localization and no mention of the gender or age of the animals examined is given. This makes difficult a comparison with our own results. Two Cestoda and one Digena recorded in this work are absent of our observations. However, several parasite taxa identified in the present work are also present in [START_REF] Malekani | Epidemiological monitoring of gastrointestinal helminthes of two wild Galagos: Galagoides demidovii and Sciurocheirus gabonensis (primates, Galagidae) originated from Democratic Republic of the Congo[END_REF]: Ascaris spp., Lemuricola spp., Ancylostoma spp., Trichostrongylus spp. and Strongyloides spp. Because of the characteristics of the cycle of the first two taxa (see discussion above), it seems likely that the parasites observed here may belong to species specific to their hosts and still undescribed.

Diversity and prevalence of gastrointestinal parasites by host species (Table 1)

Data analyses revealed the presence of nine helminth parasites and two protozoa parasites. With the exception of Lemuricola spp., present in G. demidoff only, the same parasite taxa are recorded in both host species. For both host species the arithmetic means of the prevalence values are very close: 14.7 for G. demidoff vs 15.5 for G. thomasi. However, the prevalence values observed for each parasite vary markedly. Two parasites only exhibits a high prevalence whatever which host is considered: Ascaris spp. (31.3 vs 40.6) and Balantidium spp. (19.3 vs 15.6). The differences in prevalence are particularly marked and statistically significant for Strongyloides spp. (62.7 vs 10.9) and Oesophagostomum spp. (9.6 vs 39.1).

Difference in prevalence according to collection sites and host species (Table 2)

Three parasites taxa are never represented in CDP site, whatever which host species is considered: Gongylonema spp., Lemuricola spp. and Trichostrongylus spp. For the other eight parasite taxa the prevalence is systematically higher in CDP than in DJM site, whatever which host species is considered: for G. demidoff the mean prevalence is 18.3 vs 11.9 and 20.4 vs 8.6 for G. thomasi. Proportionally, for the same parasite, the prevalence in CDP is 1,5 to 4 times its value in DJM for both host species. Finally, if the parasite diversity appears to be higher in DJM, several parasites reach their highest prevalence value in CDP; for G. demidoff: Strongyloides spp. (53.7), Ascaris spp. (46.3); for G. thomasi: Strongyloides spp. (73.7), Ascaris spp. (63.2). CDP forest is more fragmented and impacted by human activities than DJM. Previous studies have shown that these factors may influence the diversity and prevalence of parasites, particularly because forest fragmentations reduce food availability and increase host densities [START_REF] Gillespie | Forest fragmentation, the decline of an endangered primate, and changes in host-parasite interactions relative to an unfragmented forest[END_REF]. 4.3. Difference in prevalence according to the age or gender of the host species (Table 3) In our data set the effectives of males or females are quite equivalent: 79 vs 68; conversely the contrast between the representation of adults or young is high: 97 vs 50.

Gender

For both sexes the highest prevalence values deal with the same parasite taxa: Ascaris spp., Balantidium spp. and Strongyloides spp. The mean prevalence values are higher for the males than the females: 14.7 vs 11.0. However the females have a relatively stronger prevalence than the males for four parasites: Ancylostoma spp. (13.2 vs 5.1), Trichostrongylus spp. (16.2 vs 6.3), Gongylonema spp. (7.4 vs 3.8) and Entamoeba spp. (13.2 vs 3.8). Within the males the contrast between the highest vs the lowest prevalence values is higher (median = 6.3) than within the females (median = 10.3). These differences may be related with the particular behavior of the galagos where matriarchal groups are the stable units, with a family defending a territory during nomadic males are travelling over several female territories every night [START_REF] Kingdon | Kingdon Field Guide to African Mammals[END_REF].

Age

The mean prevalence values are relatively close when the adults are compared to the young: 13.0 vs 15.3; for both class the highest prevalence values deal with the same parasite taxa: Ascaris spp. (35.1 vs 36.0) and Strongyloides spp. (49.5 vs 20.0). The young have a markedly stronger prevalence (two to ten times higher) than the adults for six parasite taxa out of 11: Ancylostoma spp. (22.0 vs 2.0), Dicrocoelium spp. (12.0 vs 6.2), Entamoeba spp. (16.0 vs 4.1), Gongylonema spp. (8.0 vs 1.0), Oesophagostomum spp. (16.0 vs 3.1) and Trichostrongylus spp. (22.0 vs 5.2). However, the differences in prevalence are statistically significant (χ 2 test) only for Ancylostoma spp., Balantidium spp., Entamoeba spp. and Strongyloides spp. Within the adults the contrast between the highest vs the lowest prevalence values is higher (median = 7.2) than within the young (median = 14.0). MacIntosh et al. ( 2010) and Mbaya and Udendeye (2011) have shown the existence of a link between age and gastrointestinal parasites infestation. The fact that young are more infected could be explained by their higher susceptibility to helminthiasis due to a lack of premunity (acquired immunity through exposure). The highest values observed in adults for some parasites may be related to to the phenomenon of surinfestation (increasing population of a parasite in one host due to successive reinfestations).

Conclusion

Our work reveals very similar parasite spectrum in both host species and it is notable that all the taxa identified here were found previously in other Primate species, including Man. Because the hosts are closely related and living in sympatry a questions arise for each identified parasite taxon: is it specific for the Primates as a whole, for the Galago spp., or is it strictly host specific? To answer to this question it is needed to be able to identify precisely the parasites at the species level.

This point also deals with a hypothetical role of some parasites as zoonotic agents. Because the galagos are strictly arboreal and nocturnal and are not considered to be game animals, the contamination of Man by galagos parasite(s) or the contaminations of galagos directly by Man looks unlikely. Obviously this observation is not valid for zoonotic parasites having a ubiquitous host spectrum. However and whatever which host is considered a majority of parasites have a higher prevalence in CDP site. For this reason the influence of a closer cohabitation with Man including a fragmentation of the galagos habitats may be considered.

When compared with females, males generally exhibit higher prevalence values for 7/11 parasite taxa; males also reach the highest prevalence values observed during our study for several parasites. This may be related with the behavioral differences between galago males or females.

The young have a higher prevalence value for 6/11 parasite taxa and an equal prevalence for two of them. This may be interpreted in the context of a higher sensibility to parasitism and diseases probably partially linked with an immature immune system.

Measures to protect the environment and endangered species oblige parasitologists to change their method of study. It is no longer possible to dissect numerous animals in order to collect their internal parasites. The determination of the parasites may possibly be improved through the use of molecular analysis methods. Unfortunately these methods were not usable in the context of the present study. We therefore used available data on geographical distribution, host spectrum and host specificity of the parasitic taxa in an attempt to identify them. For these reasons, this work must be considered preliminary and, in order to try to answer to the questions it asks, additional observations are needed. This includes predominately the identification and if possible the description of the parasite taxa.

Fig. 1 .

 1 Fig. 1. Map of the study sites in South-east Gabon. Distance between S.1 Primatology Center forest (CDP) and S.2 Djoumou forest (DJM): approximately 30-40 km.

Fig. 2 .

 2 Fig. 2. Species included in the study. Host: A, Galago demidoff; B, Galago thomasi. Helminthes: C, Ancylostoma spp.; D, Ascaris spp.; E, Dicrocoelium spp.; F, Gongylonema spp.; G, Lemuricola spp.; H, Oesophagostomum spp.; I, Strongyloides spp.; J, Trichostrongylus spp.; K, Trichuris spp. Protozoa: L, Balantidium spp.; M, Entamoeba spp.

Acknowledgements

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.