Enhanced biodiversity of gut flora and feed efficiency in pond cultured tilapia under reduced frequency feeding strategies

between groups fed daily with (T1) or without (T5) fertilization throughout the trial (two-way ANOVA; P > 0.05). There was, however, a significant difference between these two treatments (T1 and T5) and the fish fed on alternate days in fertilized ponds (T2) during weeks 7 through 13 (P < 0.05), but no significant difference between these three treatments at weeks 15 and 17 (P > 0.05; Fig 1a). Weight followed similar trends, except that there was also no significant difference between these three treatments (T1, T2, and T5) during weeks 13, 15, and 17 (P > 0.05; Fig 1b). Fish that were grown in fertilized ponds without supplemental feeding (T4) showed the lowest growth among all groups. The fish produced with pond fertilization alone also had the lowest survival relative to the other groups (Table 1), whose survival rates ranged from 92–98%. The estimated feed conversion ratio (FCR) was lowest for fish fed every third day with pond fertilization (T3), followed by fish fed on alternate days with pond fertilization (T2; Table 1). Fish fed daily had the lowest feed efficiency or highest FCRs regardless of whether ponds were fertilized (T1) or not (T5) as these groups had greater feed inputs, but similar growth to those fed on alternate days with pond fertilization (T2). The economic assumption of alternate-day feeding is that it would lead to more cost-effec- tive tilapia aquaculture production, which could enhance income of small-scale farmers and consumption of nutrient rich fish. A marginal cost analysis from this study indicates feeding

on alternate days along with weekly pond fertilization (T2) gives the best return on investment (Table 2) with much of the cost reduction associated with application of half the feed in alter- nate day fed fish. Net return values were also highest for fish fed on alternate days with pond fertilization (T2), followed by those fed every third day with pond fertilization (T3; Table 2). Overall, the group fed on alternate days along with weekly pond fertilization (T2) had the high- est benefit to cost ratio (BCR) but was not significantly different from groups fed every third day or not fed supplemental feeds in fertilized ponds (T3 or T4); all three of these treatments had significantly higher BCRs than fish fed daily regardless of pond fertilization (T1 and T5). Pond water quality and plankton composition All water quality parameters among groups were well within the range for suitable growth of fish in pond culture (S2 Table) [4, 44]. Ammonia was highest in the group fed daily with pond Table 2. Economic analyses of Nile tilapia (O. niloticus) subjected to reduced-feeding regimens. Fed Daily with Fertilization (T1) Fed Alternate Day with Fertilization (T2) Fed Every Third Day with Fertilization (T3) Fertilization Only (T4) Fed Daily Only (T5) Expenditure (USD/pond)�† Fingerlings cost 11.99±0.65 11.99±0.65 11.43±2.30 11.43±2.30 12.64±0.65 Feed cost 113.88±7.20b 59.82±2.16c 29.87±7.64d - - 124.05±8.19a Lime Cost 0.58±0.03 0.58±0.03 0.55±0.11 0.55±0.11 0.61±0.03 Fertilizers cost 4.60±0.25 4.91±0.27 4.67±0.94 4.67±0.94 - -

Total Expenditure 131.05±7.98a 77.29±3.01b 46.52±10.95c 16.65±3.35d 137.30±8.73a Income�† Gross return (USD/ ha) 9,774.19±1,081.58ab 9,082.98±981.20b 5,418.46±659.15c 1,770.14±294.04d 10,235.40 ±1,063.34a Net return (USD/ha) 898.09±563.51c 3,853.63±813.67a 2,250.83±997.06b 610.42±212.27c 1,421.67±612.94c BCR (Benefit Cost Ratio) 1.10±0.06b 1.74±0.16a 1.70±0.35a 1.52±0.18a 1.16±0.07b Values are mean ± SD. Values with different letters are significantly different (P < 0.05). � USD = United States Dollar; BDT = Bangladeshi Taka. Fish were sold at $1.56 USD (120 BDT)/kg [1 BDT = $0.013 USD]. †Leasing and other operational costs such as labor were excluded from these calculations. https://doi.org/10.1371/journal.pone.0236100.t002 PLOS ONE Enhanced biodiversity of gut flora and feed efficiency in pond cultured tilapia PLOS ONE | https://doi.org/10.1371/journal.pone.0236100 July 23, 2020 8 / 23