Common carp reproduction in winter using heated water in temporal climate of Uzbekistan

. In December 2022, 10 females and 2 males of common carp (Common carp) from ponds (water temperature 10°C) were placed in RAS pools and 2 females and 2 males (control group) were placed in an open pond in Uzbeki-stan. In January, the water temperature was increased by 1°C in two days in the RAS for two weeks. Since January 20, the water temperature was 22-24°C for two weeks. The fish had weigh 2950 – 3980 (average 3235) g. From February 3th, 4 females and 2 males were injected with the pituitary gland according to the routine methods and ripe eggs and milt were obtained, ripe eggs (920 g) were incubated in Zuger apparatus. Fertility was 90%, Feb-ruary 7th, the beginning of hatching of the larvae was determined. The yield of larvae from eggs was 64%. Injected at the same time, 2 females and 2 males from the control group of carp did not produce ripe eggs and milt at all.


Introduction
In Uzbekistan, there has been a significant development of aquaculture produc-tion in recent years: if in 1994 -2009 there was produced 6,000 -9,000 tons of culture fish per year, then by 2020, after government organizational measures, completely private aquaculture brought fish production to more than 100,000 tons of fish per year ( Figure 1). The semi-intensive pond polyculture of cyprinids with the culture of warm-water fishes in 2 vegetation seasons remains the main technology. For further growth of fish production in the country, the develop-ment of intensive aquaculture technologies is necessary [1]. At the same time, technologies for complex water use at reservoirs, lakes for residual water stor-age are promising; those are systems for open fish raising. To rational fish growth in such systems, it is necessary to use the vegetation season from the middle April to the early November as much as possible [2]. A promising ap-proach would be to obtain fish reproduction in winter and fingerlings raising by April. In this case, it is possible to grow commercial fish for 1 season in open systems [3,4].
Important biological characteristics of reproductive ability of fish fe-males are absolute fecundity, relative fecundity, real fecundity of concrete fish, especially in aquaculture [5,6].
The size of the ripe eggs, as well as the size of the ovulated eggs, is an important indicator of the quality of the eggs. This size is associated with im-portant life processes during embryogenesis, larval development of fish, affects hatching of larvae, and viability of offspring [7]. Common carp (Cyprinus carpio) is native fish species for the Aral Sea basin, where Uzbekistan is situated [8]. The cultured common carp form is very popular and in demand in the local market, larger fresh fish is pre-ferred.
The purpose of this study was to assess the possibility of conducting winter (off-season) carp reproduction through the implementation of tempera-ture programs in basin water with maturing breeders

Materials and methods
Uzbekistan is located in a temperate climate, characterized by hot summers (with daytime air temperatures above 35-37°C) and relatively cold winter (with an average monthly air temperature in January of -2 --4°C). The seasonality of the temperate climate in the region is also manifested in the annual dynamics of water temperature. In the plain zone of the republic in the lentic reservoirs (lakes for residual waters storage, irrigation reservoirs, fish culture ponds), the average daily water temperature in summer reaches -25-28 o C, in winter the dai-ly average water cools down to 4-10oC. The average daily water temperature warms up above 14°C (the beginning of the growth of warm-water fish) in the first half of April, falls below 14°C (the end of the growing season) at the end of October (Table 1). That, the period of rapid growth of warm-water fish (wa-ter temperature above 20-22°C) is only about 4.5-5 months, the period of low growth rate of fish is 2-2.5 months [9]. The climatic fea-tures of the region influence the choice of aquaculture objects in Uzbekistan. The work was carried out in the winter period in 2022-2023 in an exper-imental closed module with an indoor recirculating aquaculture system water in the Institute of Zoology of the Academy of Sciences of Uzbekistan. We used 3 pools with a water capacity of 3 m3 each, where the water temperature is regu-lated.
Experimental breeders (matured fish of both sexes) were brought in from a nearby aquaculture pond hatchery, where brood stock is formed starting from fry in carp polyculture with breeders of silver carp (Hypophthalmichthys molitrix) and grass carp (Ctenopharyngodon idella). In mid-December, during the catch of growing ponds and fish transfer to wintering pond, 12 random fe-males and 4 males of common carp were taken (water temperature in ponds was 10-11 °C), the fish were transferred to experimental tanks of recirculating aquaculture system with the same temperature. The fish were stocked to the tanks separately by sexes. Water aeration was provided in the tanks.
In early January, the temperature in the tanks began to rise: one day it was raised by 1°C, then it was kept at the same level for one day, and so on. By January 18th, the water temperature reached 20°C; the fish were kept in this state for 2 weeks.
The water temperature was measured with an accuracy of 0.1 °C in the tanks, as well as in an open nearby earthen pond. In that open pond, 2 females and 2 males were kept in cages (1 m 3 ) as control group. In recirculating system yanks, when the water temperature started to rise, the fish were fed with 'Aller Aqua' feeds for table common carp culture, the ratio was 3% of the fish bio-mass per day.
The experiment was begun on February 3th. From the open pond, 2 females and 2 males of the control group were transplanted into the tanks of re-circulatin aquaculture system. On that day, 4 experimental females and 2 ex-perimental males, as well as 2 females and 2 males from the control group, were injected according to the generally accepted scheme (double pituitary in-jection for females and single injection for males). We made attempts to obtain mature sexual products on February 4th. The standard incubation of eggs was carried out in Zugger apparatuses [10,11]; the hatched larvae were kept in pools (Figure 2 The state of development of the reproductive system of females was analyzed. For this, on February 18th, in the remaining 6 females (who were not injected), the standard body length (to the end of the scale cover) (SL, cm) was measured with an accuracy of 0.1 cm, the total body weight (W, g) with an accuracy of 1 g. At autopsy, the stage of maturity of the gonads was determined on a 6-point scale, the mass of the gonads (q, g) was measured with an accuracy of 1 g [11].
A piece weighing 1 g was cut out from the middle of the gonads and fixed in a 4% formalin solution. Under laboratory conditions, the number of ripe eggs in the sample was counted, recalculated for the mass of gonads, and the individual absolute fecundity (thousand eggs) of females was determined. By dividing the absolute fecundity by the body weight of females, the individu-al relative fecundity (eggs/g of body weight) was obtained.
Ovulated eggs from each female was collected in a measuring cup and the volume of caviar obtained was recorded. 1 ml of caviar was taken with a pi-pette and fixed in a 4% formalin solution. Under laboratory conditions, the number of eggs in the sample was determined, recalculated for the total volume of eggs, and the working fecundity was determined.
The size of ripe eggs in the gonads and the size of ovulated eggs were de-termined using a apparatus "Microfot-5PO-1" for microfilm reading, in which 100 eggs were drawn in a row ( Figure 3). For each egg, two diameters were meas-ured, located at an angle of 90° to each other. The half-sum of these diameters was calculated as the size of the eggs (μm), converted into absolute values, tak-ing into account the increase..

Results and discussion
In sample there were 12 common carp females with total weight 2950-3980 (average 3235) g, and 5 males weighing 3101-3800 (3230) g.
The water temperature was kept at the level of 11 -12 °C in the tanks of recirculating system in the second half of December. For two weeks in January, the daily average water temperature was raised by 1°C in two days. From Jan-uary 20nd until the end of the experiment, the water temperature was 22-24°C. The temperature in the open pond (control group) varied from 6 to 10°C dur-ing December -the whole of January.
Dried pituitary glands of common carp were used to stimulate matura-tion. The first injection to females was carried out on February 3th at 10-00 at the rate of 0.5 mg/kg of female body weight. The second injection to females (3.5 mg/kg) was carried out 12 hours later, at the same time a single injection was given to males (0.5 mg/kg of body weight). 14 hours after the second injec-tion, ripe eggs and milt was obtained from fish of both sexes (Kamilov et al., 2003). All males had given milt; ejaculate volume was 15-20 ml from each male. Of the 4 females, 3 females had given ripe eggs, the total volume of eggs was 920 g of eggs.
The fertilized eggs were incubated in two Zugger apparatus connected to the water supply system in the recirculating water supply system. The fertiliza-tion of eggs was determined at the level of 90%. On the morning of February 7th, the beginning of hatching of the larvae was noted. Hatching took place within 20 hours. All larvae were placed in a tank with a water level of 10 cm and kept there for 3 days. The number of larvae was counted -560,000 pieces. So, the rate of larvae hatching from eggs was 64%.
It should be noted that 2 females and 2 males from the control group of common carp injected at the same time did not produce ripe gametes at all.
In sexually matured 6 female carp, which were kept in tanks, by the end of February, the gonads were at the III-IV stage of maturity. The mass of the gonads varied from 358 to 625 g. At the same time, a group of maturing oo-cytes, which were all yolk, was clearly visible in the gonads. In the device "Mi-crophot -5 PO -1" they were easy to separate on such a basis that they were completely opaque in transmitted light. The oocytes of the reserve fund (the most developed oocytes were at the stage of protoplasmic growth) were all yel-lowish translucent.
The individual absolute fecundity of females varied from 168 to 1200 thousand eggs. The individual relative fecundity of females was 99.1 -301.1 eggs/g of body weight. Yolk oocytes had a diameter of 0.6-1.2 (0.98 on aver-age) mm.
Variability in the size of ovulated eggs was noted for each of the 3 fe-males, however, this variability was not high: the differences between the smallest egg in diameter and the largest from one female differed by 5-8%. In all females, the distribution of ovulated eggs approached normal, no asym-metry in the size of the eggs was noted.
Some variability in the size of eggs between females was also revealed. The minimum individual sizes of eggs in three females varied 1.1-1.3 mm, the maximum individual sizes were 1.41-1.65 mm, the average for each female var-ied 1.49-1.51 mm.
It can be noted that the features of the reproduction of carp in the condi-tions of water bodies of Uzbekistan. Spawning of wild common carp stocks in lakes, rivers and reservoirs takes place from the last ten days of April to mid-May when the water temperature rises above 18°C. In aquaculture pond hatcheries with a cultured form of common carp, reproduction is carried out at the same time. In different hatcheries farmers use either "wild spawning" or stimulation of maturation with gonadotropic hormones with artificial fertilization and incubation of eggs in Zugger apparatus. Those, when the water warms up steadily above + 18 ° C, the winter delay in the development of the gonads ends in the carp and the maturation of the yolk oocytes resumes, the transition of the gonads to IV, and then the transition to the V stage of maturity oc-curred.
We tried to achieve the maturation (resumption of common carp matu-rated yolk eggs) by heating the water to spawning values and to stimulate mat-uration by using gonadotropic injections. The experiment showed that in such conditions all the experimental males produced milt in the winter, as did 3 females out of 4 who were injected. At the same time, we not only gradually raised the temperature of the water, but also kept the fish in it for 2 weeks.
Global cage aquaculture has developed more in northern or tropical envi-ronments with relatively constant water temperatures during the year. In Uz-bekistan, located in the southern zone of the temperate climate, the growth of warm water fish in cages will be interrupted with a seasonal drop in water temperatures. At the same time, the idea of growing marketable fish fed with high-protein aquafeeds is promising and requires an assessment of the effect of feeding to accelerate growth in addition to off-season reproduction and cultiva-tion of planting material by April. The development of intensive culture of fish and feeding them with high-protein balanced granular feed in cages in-stalled in lakes for residual waters storage and in reservoirs in the conditions of the plain zone of Uzbekistan is one of the most promising. Currently, the list of aquaculture objects is limited in the republic due to limited reproduction and production of fingerlings [7]. Carp is one of the few such species, especially since it is very popular in the local market [9]. However, cages will compete with pond carp, where fish are fed much cheaper supplemental feeds. One of the ways to increase the competi-tiveness of cages will be promising when solving the issue of growing marketa-ble fish in 1 year. Then the carp from the cages will compete with the commer-cial carp grown in ponds for 2 years. The indicated is limited by regular sea-sonal changes in the temperature regime of water.

Conclusions
To grow marketable table common carp in one vegetation season, a technological method of artificial reproduction of carp in winter is required, further rearing of larvae to fry and to fingerlings by mid-April, which can be done in conditions of closed water supply systems with constantly warm water.
As our experiment has shown, in winter in Uzbekistan it is possible to successfully reproduce common carp with the use of gonadotropic stimulation of the maturation of breeders. In our experiment, we acquired fish in December of a broodstock from a pond fish hatchery, such fish -breeders have actually already stopped maturation due to the winter cooling of the water.
An increase in water temperature in January made it possible to stimulate the maturation of the gonads, and the fish responded positively to injection with the pituitary gland. All general technological norms applied in the pond hatchery are also applicable in winter reproduction. Further development of the technology can be planned in the constant maintenance of the carp brood stock in the condi-tions of recirculating water supply installations without winter stoppage of growth.