FBIP: Stellenbosch University: Freshwater Prawns Survey

Study Extent

Eastern Cape and KwaZulu-Natal

Sampling

Sample collection and field work: The current survey project is designed to run over a two year period (2015 -2016). Given the extensive geographic scope of the project and the large number of rivers to be sampled it can only be practically feasible to undertake the survey over a two year period. During the current year, 2015 the Eastern Cape province rivers will be sampled, these include the following, Diep, Gamtoos, Krom, Bushmans, Kowie, Kap, Great Fish, Keiskamma, Laing, Nahoon Dam, Great Fish and Kei. During the following year, 2016, KwaZulu-Natal rivers will be sampled, these include the following rivers, Mthatha, Mzimvula, Mtentu, Mtamvuna, Mzimkulu, Mkomazi, Mgeni, Mvoti,uThukela, Mfolozi, Nyalazi, Msunduzo, Mkuzi and Pongola. At each site we will sample three localities (upper, middle and lower reaches) taking a minimum of ten samples per sample locality (yielding an estimated total of 780 prawn specimens). Samples will be collected, using baited traps, hand nets or trawling - depending on the depth of the river. Traps baited with fish or dog food will be left on site for 24 hours. Upon collection of samples, GPS coordinates will be taken and the specimens will be photographed using a Nikon Digital Camera. Samples will be euthanized with clove oil, and preserved in absolute ethanol in honey jars. Samples will be identified using keys together with the help of Dr De Grave, a leadings specialist on freshwater prawns. Upon completion of the sample collection and the data analyses, specimens will be deposited in the South African Natural History Museum collection (Iziko) of Cape Town. It is important to note that I have already sampled all the major rivers in the Western Cape province together with my PhD student. This work forms a natural extension of our sampling effort and all the data will be used towards her PhD thesis. (It is very important to note that we are not planning to sample the remaining provinces at this point because there is a cap on the feasibility of a survey that will include all of the remaining provinces in the country).

Method steps

1. DNA extraction: DNA will be extracted using a Machery and Nagel kit following the extraction protocol of the manufacturer. DNA will be stored in a refrigerator prior to use. We will amplify three loci. These include the barcoding locus cytochrome oxidase one (COI) and two nuclear markers, histone three and 28S. Universal primer pairs will be obtained from the literature and are currently already being used by my PhD student Ms. Wood who works on prawns in the Western Cape. PCR's will be done in a thermocycler and the PCR products will be gel purified using a Bioflux gel purification kit. Samples will then be sent to the central analytical facility at Stellenbosch University for DNA sequencing. This aspect of the study will be done after each field trip and is estimated to take about six months. All of the sequences will be deposited in GenBank / BOLD. Molecular data analyses: Sequence data will be aligned in MUSCLE and the two protein coding markers (COI and histone 3) will be checked for stop codons. It should be noted that nuclear DNA markers in Arthropods are generally slow evolving so we will explore a number of variable marker options - however, these will in the end be restricted to two loci (as per our current budget).We will collapse all the COI data into networks for each of the species using TCS. An SAMOVA and AMOVA will be performed on the COI data. We are selecting this locus because it is the most rapidly evolving locus we will use. In addition we will also construct phylogenetic trees for the haplotypes for each respective species using Bayesian inference and Maximum Likelihood. This aspect of the study is estimated to take four months. Quantification of samples for in situ fisheries: The larger Macrobrachium species have real potential for small scale in situ fisheries and we will focus on these species to estimate their abundance. Total abundance and biomass of prawns will be calculated using the swept area method. The area sampled will be calculated by multiplying the effective width of the trawl by the towing distance. To standardize data, the number of individuals or the total wet weight of the individuals captured at a sampling station will be divided by the area sampled. A mean abundance and biomass of the sampled stations will then be calculated. Abundance results will be expressed as individuals per m square, and biomass in grams wet weight per m square. Wet weight will be measured using a Sauter AR100 microbalance. This activity will happen at every sample site where Macrobrachium is collected.

Freshwater prawns

1. Decapoda
   rank: order

Eastern Cape and KwaZulu-Natal