Prevalence Of Parasites In Freshwater Fishes In The Southern Part Of Ligawasan Marsh, Philippines

Ligawasan Marsh, the largest river basin in Mindanao is home of common fishes such as dalag, pupuyo, hito, tilapia and gourami. Fish parasites pose a great threat to freshwater fishes to public health and economy of an area. This study aims to examine the Prevalence of Parasites in Freshwater Fishes in the Southern Part of Ligawasan Marsh; Endoparasites in the representative species; Mean intensity and abundance infection of the Endoparaites. The study utilized necropsy, dissection and microscopy method to examine Endoparasites. Trematodes Clinostomum sp, suspected Camallanus sp., a Neocamallanus sp. and a member of Order Spirurida, all nematodes were recovered. The study yielded that Fish parasites in Channa striata Anabas testudineus had prevalence in Kabuntalan and Datu Piang. Nine Acanthocephalan recovered in Oreochromis niloticus had also prevalence rate. Suspected trematode cysts and eggs were also recovered in Trichopodus trichopterus and Clarias macrocephalus. Thus, it is concluded that there is prevalence of nematodes and trematodes species in C. striata, A. testudineus, O. niloticus, C. macrocephalus and T. trichopterus The Camallanus sp and Neocamallanus sp, both common aquarium parasites are first found in the Southern part of Ligawasan Marsh.


Introduction
Ligawasan Marsh is located in South Central Mindanao, and is recorded as the largest swamp and marsh area on the island consisting of two adjoining marshy basins-Ligawasan and Libungan Marsh where most of the population in ARMM fish for food and source of income.
There are 344 species of freshwater fishes in the country, including the native and introduced fish species. Maguindanao had their shares in the annual increase of fish production. Nineteen point two (19.2) per cent are acquired through inland municipal fishing. Tilapia, milkfish (bangus), carp, freshwater catfish (hito), mudfish (dalag), guorami and freshwater goby (biya) were just the common freshwater species caught by inland fishermen (BAS, Phil 2012).
Local studies show the prevalence and occurrences of the endo-and ectoparasites in Ligawasan Marsh. A Study made by Salcedo et al (2009) on fishes sold at the Kabacan Market to which one of the sources is the northern part of Ligawasan Marsh. showed a frequency rate of parasitic occurrence of 1.87% (Ligawasan Marsh) and 6.97% from Pulangi River which geographically is connected to Ligawasan Marsh. Other local studies previously done on fishes examined coming from Kabacan Public Market recorded absence of trematodes and nematodes and prevalence of tapeworms (Fernando & Gonzaga 2004). Same results were recorded on the study done in Midsayap Public Market. Among the three fish species examined, cestodes (tapeworms) were the only collected fish parasites.
Recently, Fortinez, (2016) conducted a study on Kuyapon side of Ligawasan Marsh and revealed fecal contamination as well as presence of zoonotic nematodes such as Trichuris sp and Ascaris sp. A trematode species Clinostomium philippinensis and Ascaris testudines were recorded to be infesting fish species of Channa striata.
All of these studies are indicative of greater possibilities of parasite infestations in fishes in the Southern area of Ligawasan Marsh, which has been unstudied so far.
The general objective of this study was to determine the prevalence of the fish parasites in selected areas of the southern portion of Ligawasan Marsh. Specifically, this study determined prevalence of endoparasites, mean intensity of infection, and abundance of the endoparasites.
Studies on fish parasites have long been contributing worldwide to the current knowledge of the diversity and biology of the ecto-and endoparasites, protists and metazoans, of freshwater fishes. Numerous studies had already been conducted in the different lakes and bodies of water all over the Philippines, including Ligawasan Marsh. High prevalence and occurrences of parasites had been recorded in the northern part -Pagalungan side of Ligawasan marsh. However, there is no data gathered yet from the southern part of Ligawasan Marsh. In this study, the level of infestation and occurrences of parasites in the two sampling sitesmunicipalities of Kabuntalan and Datu Piang, all under ARMM were taken into account. Expectedly the outputs of this study would educate the local residents on the implications on this health presence of zoonotic parasites, in their local fish commodities, on their health. Moreover, this study also served as the basis for the Local Government of Kabuntalan and Datu Piang to come up with a resolution to preserve the natural environment of Ligawasan Marsh by educating the residents on proper waste disposal and for the Department of Health to spread awareness campaign as to the impact of parasites on health issues and importance of proper sanitary practices. Also by knowing the level of infection of the parasites on the health of the fish, something can be done to address them, thus increase the income of the fishfarming communities in that part of Ligawasan Marsh.The study also integrated new findings regarding parasites in Ligawasan Marsh with some information and references gained from prior studies in order to add to the understanding of the parasite existence as well as on its ecological relationship and importance.
This study was conducted purposely to examine and assess the existence of zoonotic endoparasites in freshwater fishes in the southern part of Ligawasan Marsh. However was only limited to the occurrence and incidence of the endoparasitic infestation in freshwater fishes on species of Channa striata (dalag), Oreochromis niloticus (tilapia), Clarias macrocephalus (hito), Trichopodus tricopterus (guorami), and Anabas testudineus (pupuyo) which are the most consumed fishes in the area.

Research Design
The research design used in the study was survey. Experimental processes were done in standard laboratory procedures.

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Locale of the Study Ligawasan Marsh is an extensive swamp region with a circumference of 25mi (40km) long and 20mi (30 km wide along the Pulangi River in South Central Mindanao, Philippines. It is a vast complex of river channels, small freshwater lakes and ponds, extensive freshwater marshes. It is potentially diversified in freshwater fishes of great economic values and importance.

Collection of Samples and Identifications
The two ARMM municipalities namely, the municipalities of Kabuntalan, and Datu Piang served as sampling sites where fish samples were collected. Fresh fish were purchased from the contracted local fishermen, on a daily basis.
The purchased live fishes were placed in storage ice chest, transported to the laboratory of the Department of Biology in University of Southern Mindanao, identified using pictorial keys, recorded length in centimeter (cm) and weighed in gram(gm), photodocumented and were processed for parasitic examination. The total purchased of each type of fishes were approximately thirty (30).

Sample Preparation, Identification and Fixation of the Parasites A. Internal Examination
Each fish examined was laid on one side and was opened from the anal region to the mouth via ventral incision. A pair of blunt/sharp scissors was used, with the blunt end going into the body cavity to minimize damage to the internal organs. The internal organs as well as the entire body cavity and intestine were inspected for the parasites. The collected suspected parasites were soaked in 9% saline solution.

B. Fixation
In this study, the parasites extracted from the fishes were fixed with 70% ethanol and/or formalin.
B.1. Staining for Nematodes The nematodes were fixed in glacial acetic acid overnight and stored in 70% ethanol with 10% glycerin. Nematode morphologies were studied in temporary mounts by then removing from alcohol and clearing with glycerin. Before returning the nematodes to storage, it was rinsed away with clearing fluid using 70% ethanol. Staining was done for detailed identification.

B.2. Staining for Trematodes and Cestodes
The trematodes and suspected trematode metacercariae were released, pressed between glass slides and stored in 70% ethanol.
Flukes (trematodes) were stained using acetocarmine staining technique. Metacercariae were stored in 70% ethanol directly. Flatworms were usually stained and mounted as permanent mounts.
B.3. Staining for Acanthocephalans Acanthocephalans were placed into 70% ethyl solution until the staining procedure. The specimen were stained using the acetocarmine staining procedure. The specimen was placed in an epindomorph tube. By pipetting solutions in and out of the vial containing the parasites, staining was accomplished.
The staining technique used was the Semichon's Acetocarmine Technique. C. Identification of Parasites The parasite identification was done using identification dichotomous keys and pictorial guides. The pre-identified parasites were then confirmed by the experts of the USM Biological Departments of the College of Arts and Sciences.
Proceding of the International Conference on Intellectuals' Global Responsibility 2020 (ICIGR): Science for Handling the Effects of Covid-19, Facing the New Normal, and Improving Public Welfare

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Proceding of the International Conference on Intellectuals' Global Responsibility 2020 (ICIGR): Science for Handling the Effects of Covid-19, Facing the New Normal, and Improving Public Welfare To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.

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Un. Trematode 2 Legend: (√) means presence of parasites and (−) negative for the parasite. Kab = Kabuntalan, DP=Datu Piang There were 7 species of parasites identified. Of them 2 were trematodes, 2 were nematodes and 1 acanthocephalan. In addition, there were 3 unidentified parasites including suspected trematode, suspected nematode and some suspected parasite eggs. Table 2 shows parasite prevalence in both Kabuntalan and Datu Piang sites. Prevalence is the percentile representation of infected hosts divided by hosts examined multiplied by 100.  Abundance of parasites are determined by the number of parasites collected in a sample over the total number of fish examined. Table 3 illustrates the high abundance rate of C. striata in terms of parasites collected in Datu Piang site, followed by T. trichopterus and A. testudineus. There was a tremendous drop in the graph as it illustrates the lowest abundance in C. macrocephalus as well as the absence of collected parasites in the T. trichopodus and O. niloticus in C. punctatus in Kabuntalan site.

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In the study of Miah, et al., 2013, C. punctatus was infected by 7 species of parasites. Of these, four were trematodes ( Genarchlopsis bangladnesis, Allogamtitrema attu, Phyllodistomum sp., Neoppecocline saharanpuriensis); two were nematodes (Ascaridia sp., Procamallanus sp.) and one was an Acanthocephalan (Pallisentis nandai). The findings of this study is almost the same with the findings of Miah, et al., 2013. Furthermore, in this study there were five trematodes: Clinostomum sp., Posthodiplostomumsp. and three unidentified trematodes; three nematodes: Camallanus sp and Neocamallanus sp. and a member of Order Spirurida as well as 1 Acanthocephalan were found. But, the trematodes and nematodes found in C. striata were also abundant in A. testudineus of this study and Order Spirurida sp was only found in the A. testudineus. In addition, unidentified trematodes of Clinostomum sp. and E. heterostomum resemble those recovered in T. trichopterus but Acanthocephalan sp. were only present in O. niloticus and were not found in C. striata and A. testudineus. However the dissimilarities in fish hosts of the mentioned trematodes, nematodes and acanthocephalan in both studies may be attributed to the factors such as feeding habits of the hosts, water quality in terms of habitat and availability of infected intermediate hosts (Kundu & Bhuiyan et al., 2016).
The findings of this study is consistent with the previous studies, that recovered common intestinal nematode Camallanus anabantis from A. testudineus (Mandal et al., 2016). The intestinal parasites was also reported to be harbored by many other indigenous fishes such as C. striata and T. trichoptrus and the Puntius sp. only caught in Datu Piang site.
The relative density of infection (Table 4) derived from the number of collected parasites in a sample over the total host examined reflects a higher account on T. trichopterus in Datu Piang site followed by A. testudineus, still in Datu Piang site. All in all, the Datu Piang In the study of Dhole et al. in 2010, Mastacembalus armatus a fish species in India, constantly showed higher parasitic infection. In comparison, the fish they used, and the T. trichopterus used in the present study, were heavily infected with parasites in one site but showed absence of parasites in the other site. Moreover, in this study the intensity of infection of parasites was so high in the fishes caught in the Datu Piang site. Some workers concluded that fish from more polluted waters tend to harbor more parasites than from less polluted area. Looking at the areas of Kabuntalan and Datu Piang in the map (Fig. 17) Barangay Kalipapa (yellow) in Datu Piang has lesser area and the risk to pollution is higher than Brgy Butiren (pink) in Kabuntalan. Furthermore, according to Dhole' et al., 2010 as cited from Polanski (1961a) the main factors determining the variety of parasite fauna as well as the intensity and incidence of infection can be summarized as follows: the diet of the host, life span of the host, the mobility of the host throughout its life including the variety of habitat it encounters, it's population density and the size attained, where large host provides more habitats suitable for parasites than small ones. The last statement though was contradictory to the findings of this study where prevalence, intensity and abundance of parasites recorded were high in A. testudineus and T. trichopterus which were low C. striata and C. macrocephalus. Figure 16 shows that intensity of parasitic infection is consistently higher in Datu Piang in all fish species than intensity of infection in Kabuntalan. Higher infection rate must be attributed to the time and seasonal factors during the time of catching. In the study made by Mgwede and

Conclusion
A total of one hundred fifty (150) fish samples were examined, from five common fish species, namely Channa striata, Anabas testudineus, Oreochromis niloticus Clarias macrocephalus and Trichopodus trichopterus.
The two municipal sites of Ligawasan Marsh -Kabuntalan and Datu Piang were found positive for the presence of trematodes such as Clinostomum sp., Posthodiplostomum sp. and three unidentified trematodes. Nematodes were also present in both sites such as Camallanus sp. and Neocamallanus sp., and a member of Order Spirurida. There were numerous unidentified eggs recovered in both sites, and an Acanthocephalan in the Datu Piang.
It can then be concluded that there is prevalence of nematodes and trematodes species in C. striata, A. testudineus, O. niloticus, C. macrocephalus and T. trichopterus The Camallanus sp and Neocamallanus sp, both common aquarium parasites are believed to be the first to be found in the Southern part of Ligawasan Marsh.  To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.