Some biological aspects and molecular variations in frigate tuna, Auxis thazard of the coastal waters around Sri Lanka

of Sri Lanka. For this period, the length-weight relationship and Fulton’s condition factor (K) calculated for using standard length measured in centimetres and weight measured in grams were shown as = 0.1091 3.3385 and 1.93, respectively, indicating a relatively healthy growth pattern and a comparatively unpolluted habitat. Studies on the stomach contents revealed that are non-selective carnivors,


INTRODUCTION
(frigate tuna), also called in the local market, is a very important neritic tuna species in Sri Lanka. There are three species of neritic tuna, (kawakawa), (frigate tuna) and (bullet tuna), commonly occurring in the coastal waters of Sri Lanka. The neritic tunas constitute approximately 13 % of the total tuna production of the country (Bandaranayake & Maldeniya, 2012). In 2015, has contributed 38 % while and have contributed 39 % and 23 %, respectively to the total neritic tuna catch of Sri Lanka (Rathnasuriya et al., 2017). A higher percentage of neritic tuna production has been recorded from the Southern and et al., 2014). It has been reported that in the recent past, Sri Lanka, India, Indonesia and Iran together have accounted for 90 % of catch of the world (IOTC, 2014). Frequently, and the catch is reported as a combination of these two species. Neritic tunas are caught by a variety of gear types, such as gillnets, handline and troll-line in the coastal waters around the country (IOTC, 2016).
However, there is very limited information available on the biology and stock structure of of the Indian Ocean (IOTC, 2016). The biology, genetics have been studied in India (Ghosh et al., 2012;Kumar et al., 2012a) and Indonesia (Noegroho et al., 2013;Hamidi et al., 2018 Although a few biological studies have been carried out in Sri Lanka on , genetic studies have not yet been carried out. Hence, studying the biology, reproduction and genetic stock structure of is very important as this information can be compared with the information available for the region to manage the

Sample collection
A total of 405 from the coastal areas of Chilaw and Kalpitiya in the was carried out from October 2015 to September 2017 from day-boats, which had gone to sea in the evening and returned the next day morning.

Morphological and biological analysis of A. thazard
Biological analysis was carried out for the 405 locations. Weight to the nearest 0.1 g and standard length relationship between standard length and weight of the ... (1) where, is the standard length in cm, a is the regression intercept and b vs log L (Froese, 2006).
Fulton's condition factor (K) was estimated from the following relationship: ... (2) where, is the standard length in cm and is the weight in g (Sarkar et al., 2013).

Feeding and spawning of A. thazard
The stomachs were weighed to the nearest 0.1 g. The stomachs were then cut open and according to the stomach fullness observed visually, it was categorised into 5 classes: full, three fourths (3/4) full, half (1/2) full, one fourth (1/4) full and empty. The prey items were found within the stomach cavities. The weight of each category of prey item was recorded to the nearest 0.1 g (Hyslop, 1980). The gonads were analysed to determine 1974) given in Table 1. The weight of the gonads and maturity stages were recorded. Fecundity was calculated by counting the number of eggs in a known quantity of preserved mature stage IV female gonads. Gonado somatic index (GSI) was calculated for male and female samples according to the following equation (Adebiyi, 2013): ... (3) (gonad weight + stomach weight)] weighed in g.

Molecular analysis of A. thazard
Tissue samples of 75 randomly selected samples collected from all the locations were used for total genomic DNA extraction using a standard phenol chloroform method (Green & Sambrook, 2012 of the extracted genomic DNA, 2.5 units of Taq MgCl 2 . Forward and reverse sequences were obtained for each sample and the consensus sequences were derived by using BioEdit version 7.0.5.3. These likelihood tree was created for 75 of these sequences using the Mega 7 software programme using a bootstrap populations.

Length weight relationship and Fulton's condition factor K for A. thazard
Length and weight parameters were measured for a total of 405 the locations. highest mean weight and standard length was recorded from the Southern province and the highest K value was from the Western province. The standard length of the collected samples varied from 19.4 to 40.7 cm and the weight of the samples varied from 118.9 to 1430.9 g. The congruence with the length range of caught in the Indian Ocean, which has been reported as 20 cm -50 cm (IOTC, 2016). The length weight relationship was represented by the equation = 0.1091 3.3385 obtained according to the LW relationship ( Figure 2). because it can be used in the estimation of growth with increases in length. The b value of the length-weight relationship equation is used to determine the type of somatic growth. The b value being 3.3385 in this study, showing a positive allometric growth pattern (Froese, 2006).
The Fulton's condition factor K for the sampled was 1.93. The K factor can be used for understanding the 2002). A K value of more than 1 indicates a healthy condition while a value less than 1 indicates a poor condition (Hall & Van Den Avylle, 1986). In general, the higher the K value, the better is the physical condition value obtained for condition is relatively good.

Feeding and spawning of A. thazard
Feeding was evaluated according to Hyslop (1980) by the frequency of occurrence method. The percentage of each group of prey item was calculated to show of stomachs were full and 19 %, 17 %, 24 %, 9 % and 6 % were three quarters full, half full, quarter full, one eighth full and empty respectively (Figure 3). This revealed 90 % of them were at least one eighth full. To study the food habits and feeding habits of , the stomach contents were examined. The results showed the presence of diverse prey items such as shrimps, indicating that is a nonselective feeder anything that is abundantly found in the surrounding environment. Further, the highest percentage (47.8 %) of prey items was crustaceans, including shrimps, crabs and stomatopod larvae (Figure 4). The next most of species such as , , and content. Fish and crustaceans together could be seen in could be seen in 1.9 % of the stomachs. In addition to and 3 stomachs (1.9 %) contained plastic and polythene.
A major proportion of all the stomachs analysed (55.4 %) contained totally digested food material, which could not studied from the Sri Lankan waters are in accordance with the feeding patterns of caught along the Indian coast (Rohit et al., 2017).
The gonads were analysed to determine the in Table 1. the GSI peaked during the period May to July ( Figure 6) and in males from May to August, indicating that their spawning season is during this period. The ovaries of mature females carrying eggs increase in size just before spawning and the GSI value decreases after spawning.
The highest GSI values of male and female being during the period May and August indicates that the species' spawning period is from May to August in the waters around Sri Lanka. However, IOTC (2016) has reported that the spawning period for the northern Indian Ocean frigate tuna species is from January to April. This deviation could be due to the variation in temperatures of The fecundity of female was shown to be 48,056 to 267,000 eggs. These results show that the fecundity of is lower than shown in previous studies (Tampubolon et al., 2016), indicating that there could be variations in the nutritional status, salinity or the

Molecular analysis of A. thazard
The mitochondrial D-loop region of 75 randomly selected samples collected from the coastal waters of the four provinces of Sri Lanka were sequenced (GenBank accession numbers MH557799 to MH557821). The blasting of these sequences using the NCBI blast facility resulted in identity of the species with more than 97 % similarity.
These sequences were used for the maximum likelihood tree construction ( Figure 8) and the results regions are clustering together and can be considered as originating from one common stock.
The knowledge about stock structure is very it is assumed that the stocks being managed comprise of a single stock. This is due to the reproductive potential, mortality and adaptations to exploitation, changing et al., 2012b). The phylogenetic analysis of the 75 sequences being mixed within clusters indicates that the stocks of common stock, indicating that they can all be managed as one stock in the future.

Mean of weight
Mean of standard K factor Table 2: Mean values (mean ± SD) of weight, standard length and K factor for provinces

CONCLUSIONS
The frigate tuna found in the Sri Lankan coastal areas have a healthy growth pattern and live in an environment that is favourable for their growth. They are non-selective carnivorous feeders, feeding on a range of prey items available in the environment. The spawning season of in the Sri Lankan coastal areas is from May to August. Molecular analysis revealed that individuals of country appear to belong to the same stock.