Zooplankton Dynamics of the Kienke Estuary (Kribi, South Region of Cameroon): Importance of Physico-Chemical Parameters

All Cameroonian estuarine systems, like the Kienke estuarine system (urban area of the port city of Kribi), are considered, as everywhere in the world, as unstable and vulnerable coastal ecosystems insofar as they are influenced by anthropogenic activities (port facilities, industrial facilities), without forget-ting climate change. The present work was initiated in order to assess the influence of the seasonal evolution of physico-chemical parameters on the dynamics of zooplankton in the estuarine system of the Kienke. A study to assess the influence of seasonal evolution of some physico-chemical parameters on Zooplankton population dynamics was conducted from June 2016 to August 2017 in the Kienke estuarine system (Kribi, South Cameroon Region). Samples were collected in five (05) sampling points at the lower stream, at the confluence and then at 100 meters from the bank at sea following a monthly frequency. The Kienke estuary was characterized by spatio-temporal variations of physico-chemical parameters. These parameters are high temperature, relatively high electrical conductivity and salinity, and a relatively basic hydrogen potential (pH). Nutrients (ammonia nitrogen, nitrates and orthophosphates) were relatively low in the Kienke estuary. The organic pollution index (OPI) indicated moderate to high water pollution. At the surface and at depth, during the long dry season (December to February), Zooplankton densities were very low in the Kienke estuarine system. But rather high during the main rainy season (August to October). The results show that 105 species of Zooplankton belonging to 46 families grouped into four orders were identified. At the surface, 52 species of Zooplankton belonging to 23 families and 4 orders were identified, while at depth, 53 species of Zooplankton belonging to 23 families were also identified. The most abundant group was the Copepods represented by the following species: Tropocyclops confinis Kiefer, 1930; Mesocyclops sp. Sars, 1914; Macrocyclops sp. Claus, 1893; Thermocyclops sp. Kiefer, 1929; Parvocalaus elegans Adronov, 1972 and Clausocalanus sp. Giesbrecht, 1888. Overall, there was a predominance of microcrustaceans (Cladocera and Copepoda) over rotifers. The results obtained in this work will be of capital importance for the elaboration of sustainable management policies for the estuary of the city of Kribi.


Introduction
Estuaries are privileged areas for human activities. Indeed, these transition zones between continental and marine waters are favorable places for the development of economic activity [1]. Estuaries are also sites of great biological interest in which physical, chemical and biological interactions generate ecosystems among the most active of all natural environments. The interest of estuarine ecosystems for marine species is linked to the presence of high populations of Zooplankton, essential links in the trophic chain between primary and secondary production, which make estuaries ideal nursery areas for the development of larvae and juveniles of crustaceans and fish (estuarine ecophase). Estuaries are also sites of great biological interest in which physical, chemical and biological interactions generate some of the most active ecosystems of all natural environments. They constitute a gateway for the transport of organic matter of continental and oceanic origin, either from upstream to downstream, or from downstream to upstream depending on the conditions, the rhythm of the tides and the seasonal fluctuations of the flows [2]. The interest of estuarine ecosystems for marine species is linked to the presence of high populations of Zooplankton, essential links in the trophic chain between primary and secondary production, which make estuaries ideal nursery areas for the development of larvae and juveniles of crustaceans and fish (estuarine ecophase) [3]. Thus, the different species of the estuarine pelagic ecosystem have developed strategies to migrate or maintain themselves in these environments favorable to their growth and/or reproduction [4]. One of the key phenomena generated by the interaction of stream dynamics and tidal dynamics, especially in high tidal seas, is the formation of a zone of maximum turbidity [5]. Previous studies have reported the practice of industrial fishing in these areas since 1912 [6]. Crosnier [7]; Raitt and Niven [8] observed the daily behavior of shrimp on experimental trawlers to guide fishermen. Le

Study Site Parameters
The study was carried out from

Presentation of the Sampling Points of the Kienke Estuary
The sample points codes, geographic coordinates, altitudes and main activities of these sample points are represented in Table 1. Overall, it appears that this estuary is located between 9˚53'6'' and 9˚55'48'' East latitude and 2˚55'30'' and 2˚57'54'' North longitude with an average altitude of around 10 m above sea level and an economic activity that revolves around artisanal fishing ( Figure 1).
Tous les systèmes estuariens Camerounais à l'instar, du système estuarien de la Kienke (zone urbaine de la ville portuaire de Kribi) sont tous considérés partout dans le monde, comme étant des écosystèmes côtiers instables et vulnérables dans la mesure où ils sont influencés par des activités anthropiques (installations portuaires, installations industrielles) sans toutefois oublier le changement climatique. C'est dans ce contexte que le présent travail a été initié afin de comprendre l'influence de l'évolution saisonnière des paramètres physico-chimiques sur la dynamique du zooplancton dans ce système estuarien de la Kienke.   All Cameroonian estuarine systems, like the Kienke estuarine system (urban area of the port city of Kribi), are considered, as everywhere in the world, to be unstable and vulnerable coastal ecosystems insofar as they are influenced by anthropic activities (port installations, industrial installations), without forgetting air conditioning. It is in this context that the present work was initiated in order to understand the influence of the seasonal evolution of physico-chemical parameters on the dynamics of zooplankton in the estuarine system of the Kienke.

Sampling Water for Physico-Chemical Analysis
Shifty on the Kienke stream was carried out using a 15 horsepower outboard motor boat (YAMAHA Enduro). Samples destined for physico-chemical analysis were collected at three sampling points at the level of lower of the Kienke watercourse, then at the level of the confluence and finally in open water (sea) 100 m from the river bank. For each station 250 and 1000 ml of water samples were collected in double-capped polyethylene containers and transported to the Laboratory of Hydrobiology and Environment in a cooler containing carboglaces for laboratory analysis APHA [16] and Rodier et al. [17]. These samples were collected at the surface.

Zooplankton Sampling
Biological samples were collected from the surface precisely at the level of the lower course (lentic area), using a 10 L bucket after having stirred the herbarium and filtered through a 64 µm sieve of 10cm in diameter. At the estuary, the filtration was carried out in a longitudinal radial and vertical manner using 200 µm plankton net. The process was repeated ten times to achieve a volume of 100 ml, for the surface samples; the water sample was taken from the quiet areas of each sampling points. While for the depth samples, the samples were collected using the six (6) liter Van Dorn Bottle and then poured into water. The collected filtrate of each sample was introduced into a 0.25 L test tube, of which 0.1 L (not fixed) was used for observations on living organisms and 0.15 L fixed with 0.01 L of formalin (5%) was used for identification and counting [18].

Organic Pollution Index (OPI)
The Organic Pollution Index (OPI) was calculated. This index was calculated from the quality classes obtained for the concentrations of the three variables; Ammoniacal Nitrogen (

Evaluation of the Zooplankton Diversity of the Sampled Waters
A qualitative and quantitative study was carried out on the Zooplankton.

Qualitative Analysis
In the laboratory, the samples were homogenized by shaking. Using a pipette, 10 ml of the sample was taken and poured into a 90 mm diameter grid Petri dish. Using a binocular magnifying glass of the brand WILD M5 at 25 x and 50 x magnifications, the identification of Zooplankton species was carried out using specific keys and works of Amoros [19]; Zebaze Togouet [20] and Fernando [21]. The identification of Rotifers was referred to the keys and identification works of Koste [22]; Durand [23]; Pourriot and Francez [24]; Dussart and Defaye [25]. The organisms, which could not be identified with a binocular magnifying glass, were mounted between slide and coverslip for observation under the IVYMEN brand microscope in order to identify them down to the scale of the species. Regarding Cladocerans, their identification is based on the observation of morphological characters, such as the shape of the body, the shape of the cephalic capsule in ventral or dorsal view, and the detailed examination of the appendages of the post-abdomen. This identification was done with a WILD M5 binocular magnifying glass after dissection, using the keys and identification works of Dumont [26]; Al-Yamani and Pruso [27]; Sharma and Sharma [28]. As for the Copepods, they are identified on the basis of the shape of the body, the length of the antennules and antennae, the lateral ornamentation of the segments of the abdomen, the position of the ovigerous sacs, the number of eggs in the ovigerous sacs and the shape of the rostrum. This identification was done with a WILD M5 binocular magnifying glass after dissection, using the keys and identification works of Leszek and Rybak [29] Al-Yamani and Kolesnikova [30]; Beleem and Kamboj [31]; Juan and Tores [32]; Jaume and Lopez [33].

Quantitative Analysis
Counting of individuals was done on the fixed sample. In fact, 10 ml of homogenized sample was taken with a calibrated pipette and introduced into a grid Petri dish 30 mm in diameter in which the counts were carried out on five samples of 10 ml each time. The density was calculated as follows: D = (nv)/V (2) where D is the density (expressed in individuals per liter), n is the number of individuals found in the volume of water analyzed under the microscope, v is the volume of water analyzed (ml) and V is the volume of filtered water (ml). The results obtained were used to calculate various indices making it possible to characterize the composition and evolution of Zooplankton.

Specific Richness
The specific richness of an ecosystem is the number of species found regardless of the number of individuals each taxon represents. This can only be assessed through a sample and for this reason it may differ from actual richness. The specific richness may well be a distinctive criterion of the ecosystems or of the sample points studied within a given ecosystem. This measure has the advantage of allowing an initial assessment of the richness of the environment from a qualitative point of view. This richness depends on the volume of water withdrawn. This is why it is important to keep a similar sampling method when surveying  [34]. The Shannon and Weaver Index represent a richness of information about the stand structure of a given sample and how individuals are distributed among different species. A low richness index indicates that the community is young with high multiplication power with dominance of one or a few species, while a high index characterizes mature populations with a complex specific composition with relatively high stand stability of Iltis [35].

Index of Diversity
The diversity index chosen is that of Shannon and Weaver [36] because it accounts for the diversity of the species that make up the stands in an environment. It establishes the link between the number of species and the number of individuals in the same ecosystem or in the same Community [37].

Statistical Analysis
The Analysis of Variance (ANOVA) and Student Fischer tests made it possible to compare biological indicators (specific wealth, abundance, diversity indices and Zooplankton biomasses) in time and space [39]. It makes it possible to assess the level of dependence between the different variables in the same ecosystem. Thus, Spearman's correlations were sought between the physico-chemical variables and the biological variables [40]. All of these tests will be performed using Statistical Parkage for Social Sciences (SPSS 20.0) software. In this study, Principal Component Analysis (PCA) was used to establish the abiotic typology of sampling points based on all of the environmental parameters measured at each sampling points throughout the study. The objective of this descriptive factorial statistics method is to present in graphic form the maximum amount of information contained in a large data table [41]. The principal components are obtained by the diagonalization of a matrix which, depending on the nature of the initial variables, is either the correlation matrix or the covariance matrix [42]. The correlation matrix was used. There are two types of representation; the scatterplot of the variables which is a correlation circle; and the scatterplot of the sites. The initial percentage explained by each principal component is shown in the form of a histogram. XLSTAT version 11.0 software was used for this analysis.
The abiotic typology of the different sampling points was made by Discriminant Factor Analysis (DFA) in order to highlight the parameters discriminating.
The Discriminant Factor Analysis (DFA) approach consists in producing a series  [43]; Desbois [44]; Villanueva [45]. The Monte Carlo permutation test (n = 1000 random permutations) was performed in order to assess the reliability of the Discriminant Factor Analysis (DFA) [46]. The Discriminant Factor Analysis (DFA) was done using XLSTAT software version 11.0.

Physico-Chemical Variables
The surface water temperature of the Kienke estuary varies between 23˚C and

Organic Pollution Index (OPI)
The Organic Pollution Index is 1.75 in the Kienke estuary in March, where organic pollution is very high (Table 3). But in the months of May, June, July, August and September the Organic Pollution Index is 2.5. The Organic Pollution Index varied between 3 and 3.25 respectively in November, February and January in the Kienke estuary where organic pollution is moderate. Organic pollution    values measured from one season to the next in the Kienke estuary generally indicated that organic pollution is strong and even very strong during the four seasons studied during our study ( Table 3). The average Organic Pollution Index (OPI) calculated from one season to another in the Kienke estuary has indicated that the waters are loaded with organic matter during the long rainy season (2.75) followed by the long dry season (2.75), the short dry season (2.25) and the short rainy season (2.25).

Principal Component Analysis of Physico-Chemical Variables
A Principal Component Analysis (PCA) carried out using the values of the 13 physico-chemical variables measured at the 5 study sampling stations shows that the first two axes F1 and F2 explain 81.12% of the information. The F1 axis of the Principal Component Analysis (PCA) explains 57.10% of the total variance. It is positively and strongly correlated with nitrate ions, electrical conductivity, dissolved oxygen, Biological Oxygen Demand, which is opposed to turbidity, nitrate ions, ammoniacal nitrogen, phosphate ions, (Figure 5(a)). This axis characterizes water, turbid, colored, rich in organic matter and dissolved oxygen because the more the organic matter is important, the more the characteristics of these variables are important. We can therefore think that this axis indicates the quality of the water, that is to say the degree of organic pollution. The F2 axis (24.02% inertia) indicates temperature and hydrogen potential. These axes combine at the same time, but with a dominance, the physical characteristics of the waters. This is the axis of mineralization and physical pollution. These two axes made it possible to divide the study sampling points into 2 groups ( Figure 5 Figure 6).

Quantitative Aspect
Quantitatively, Copepods were highly dominant in the Kienke estuary studied and constituted 76.75% of total abundance. They are followed by Cladocerans, which respectively represent 14.09% of the total abundance of Zooplankton in the waters of the Kienke estuary. Rotifers and Ostracods were the least represented with 4.94% and 4.24% of total abundance (Figure 7).     In the depth Kienke, the dominance of Copepods over Cladocerans, Rotifers and Ostracods is very pronounced as in the depth of Kienke. Indeed, these represent 86.76% of the individuals collected in the Kienke. The Cladocerans and the Rotifers then the Ostracods present respective relative abundances of 18.85% and 6.50%, 5.46%. At each sampling points, Copepods also take dominated over Rotifers and Cladocerans and Ostracods. They constitute 73, 17%, 73.77% and 75.78%, 86.76% then 82.20% of the individuals enumerated respectively in the sampling points (Figure 8(c)). In depth, the variation in Zooplank-  (Figure 8(d)).  (Figure 9(b)).

Sörensen Similarity Index
The rates of taxonomic resemblance between the Zooplanktonic populations collected in the different sampling points. Likewise, the taxa listed at the surface of the Kienke estuary at sampling points K2 and K3 have a similarity rate of 85%; sampling points K3 and K4 have a similarity rate of 91%. The taxa listed at the surface of the Kienke estuary at sampling points K3 and K5 have a similarity rate of 88%; sampling points K4 and K5 have a similarity rate of 86%. The taxa collected at sampling points K3, K4 and K5 show high rates of similarity, with values between 91% (K3 and K4) and 94% (K4 and K5). In addition, the Zooplankton inventoried in the other sampling points is dissimilar to those obtained in the other sampling points located in estuarine zones because the similarity rates obtained are overall less than 42%.

Physico-Chemical Variables at the Abundance of Zooplankton
Organisms Spearman's rank correlations between the abundance of Zooplankton organisms and the values of physico-chemical variables revealed some significant and positive correlation. The main ones are represented in Table 5. It emerges from this table that the organisms of the indicator families of environments rich in organic matter (Sididae, Oncaeilidae, Tordaniidae, Cyclopidae) are positively and significantly correlated with the variables. Orthophosphates, ammoniacal nitrogen, Suspended Solids and electrical conductivity also showed positive and very significant correlations with organisms from these three families (Pontellidae, Lecanidae, Oncaeidae). We also note positive and significant correlations between the high values of turbidity, nitrates and organisms of the families (Testudinellidae, Moinidae, Euterpinidae, Sididae, Ectinosamatidae, Tordaniidae, Sapphinnidae, Brachionidae, Corycaeidae, Cyclopidae). The values of Color, Hydrogen potential (pH), Dissolved Oxygen (O 2 ), on the other hand, show negative and significant correlations with Philodinidae, Euterpinidae, Sapphirinidae, Oncaeidae, Testudinellidae, Lecanidae, Clytemmestidae, Parvocalanidae and Corycaeidae.
Principal Component Analysis (PCA) of Zooplankton communities in the estuaries studied. Principal component analysis was performed using the numbers of constant Zooplankton species. The objective of these analyses was to evaluate the amount of Zooplankton per sampling points and analyzed the temporal distribution of the Zooplankton community throughout study. A first Principal Component Analysis (PCA) carried out from the abundances of constant Zooplankton species collected at different study sampling points made it

Physico-Chemical
During this study, the physico-chemical quality of surface water generally varied significantly in the Kienke estuary from one study sampling points and from one sampling period to another. In the water of the Kienke estuary on the surface, temperatures are between 23˚C and 35˚C respectively in sampling points K2 and  could also be due to settling phenomena which results in the progressive deposition of solid loads during transport [52]. The high solids content could also be explained by intense erosion of the watershed, following brutal rains, such as the Moulouya Oued in eastern Morocco [53].
The Hydrogen potential (pH) values recorded during the study period (5.08 -10.78 UC) in the Kienke estuary show overall that the waters of these two estuaries are slightly basic. This basicity would be due to the exogenous contribu-  being copepods with affinity with neric waters [60]. This could be explained by the fact that the latter would have migrated from the waters of the Atlantic Ocean via the strong tides to these sampling points.
During the study period, in the waters of the Kienke estuary in depth 53 species of Zooplankton including 43 belonging to the group of microcrustaceans (Copepods and Cladocerans), 09 species to that of rotifers and 01 species of Ostracod were identified. In this group, the Copepods represent approximately (78.72%) of this abundance followed by the Cladocerans which represent (14.58%) then the Ostracods with (3.87%) and finally the Rotifers with (2.83%). Copepods were more present at sampling point KP5 in March, June and July 2017 then at sampling point KP3 in December 2016. Of the four Zooplankton groups collected, there is a predominance of Copepods. This would be explained by the fact that some species of this group have the possibility of surviving in the state of resting stages, thus allowing them to be transported from one environment to another, and thus to have a larger range of Khalki and Moncef [61]. Moreover, the positive and significant correlations obtained between the dominant species were: Tropocyclops confinis Kiefer, 1930 [63]. Pearson's chi-square (Chi2) = 4.714, p-value = 0.695 > 0.05 thus, in the Kienke specific richness does not depend on depth. There is no significant difference between the surface and the bottom at the 5% level.

Spatial Dynamics of Zooplankton Communities
The

Conclusion
At the end of this study, the general objective of which was to assess the diversity and structure of Zooplankton communities in relation to the physico-chemical quality of the water of the Kienke estuary, it emerges that the water of the Kienke estuary is relatively basic; with more or less high temperature and high Electrical In addition, Zooplankton analysis shows the dominance of Copepods over Cladocerans and Rotifer. In the future, we plan to extern the study in other areas and to reinforce database that will serve as a reference for scientific community and also consider fish and crustaceans because they feed on Zooplankton and have a great capacity to accumulate polluants. They are also good indicators of pollution.