Vol. 141: 47–52, 2020 DISEASES OF AQUATIC ORGANISMS Published online September 17 https://doi.org/10.3354/dao03515 Dis Aquat Org

NOTE Negative impact on the reproductive potential of blue jack picturatus by Kudoa infection of the ovary

A. Neves1,2,*, I. Gordo3, V. Sequeira1,2, A. R. Vieira1,2, E. Silva1, F. Silva1,2, A. M. Duarte2,4, S. Mendes2,4, R. Ganhão2,4, M. C. Peleteiro5, C. Assis1,2, R. Rebelo1,6, M. F. Magalhães1,6, M. M. Gil2,4, L. S. Gordo1,2

1Faculdade de Ciências, Universidade de Lisboa, Bloco C2, Campo Grande, 1749-016 Lisboa, Portugal 2MARE − Marine and Environmental Sciences Centre, Rua Ernesto de Vasconcelos, Campo Grande, 1749-016 Lisboa, Portugal 3Pride Veterinary Centre, Riverside Rd, Derby DE24 8HX, UK 4Instituto Politécnico de Leiria, Edifício Cetemares, Av. Porto de Pesca, 2520-630 Peniche, Portugal 5CIISA, Interdisciplinary Centre of Research in Health, Faculdade de Medicina Veterinária, Universidade de Lisboa, Av. da Universidade Técnica, 1300-477 Lisboa, Portugal 6CE3C − Centre for Ecology, Evolution and Environmental Changes, Rua Ernesto de Vasconcelos, Campo Grande, 1749-016 Lisboa, Portugal

ABSTRACT: Reproduction of Trachurus picturatus off the western Portuguese coast was studied over 1 yr. During histological analyses, the presence of Kudoa sp. was detected in advanced vitel- logenic oocytes. Kudoa infections are known to cause economic loss through the induction of post- mortem liquefaction of fish muscles, but ovarian infection as reported in this study will seriously affect the reproductive potential of the species and thus impact fisheries productivity. Only females showed gonad infection which led to total degradation of advanced vitellogenic oocytes. No macroscopic, somatic or condition indices revealed differences between infected and unin- fected females, rendering this infection event a concealed suppression of reproduction.

KEY WORDS: Kudoa · Stock reproductive potential · Suppressed reproduction · Ovary degradation

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1. INTRODUCTION coast, although highly variable, reached an average value of ca. 2800 t yr−1 in the last decade. Despite its Blue jack mackerel Trachurus picturatus is an growing economic importance, there is little informa- oceanic pelagic species found between 100 and tion on the reproductive strategy of the blue jack 575 m depth (Menezes et al. 2006) in the , apart from data on size at maturity and Ocean. In the Eastern Atlantic, it occurs from the Bay spawning season for the Portuguese continental of Biscay (France) southward to Tristan da Cunha coast (Costa 2019), Azores (Garcia et al. 2015) and and Gough Islands (Smith-Vaniz 1986). This species Canary Islands (Jurado-Ruzafa & Santamaría 2013), is commercially exploited in the southern Northeast- and the fecundity type for Madeira Island (Vasconce- ern Atlantic. Landings on the mainland Portuguese los et al. 2017).

*Corresponding author: [email protected] © Inter-Research 2020 · www.int-res.com 48 Dis Aquat Org 141: 47–52, 2020

Traditionally, reproductive potential has been ton’s condition factor (K = 100 × EW / TL3) were esti- measured by the spawning stock biomass (SSB), mated and significant differences (α = 0.05) between assuming that the number of eggs each female infected and uninfected individuals were evaluated releases is a direct function of its weight and that all with permutational multivariate analysis of variance eggs have an equal probability of survival (Myers et (PERMANOVA, Anderson 2001), using the package al. 1994). The term stock reproductive potential vegan 2.5-4 (Oksanen et al. 2014) in RStudio (RStu- (SRP) was later introduced by Trippel (1999) to more dio Team 2019). accurately represent the annual variation in egg pro- The morphology of Kudoa sp. spores was evalu- duction, being a function of spawners’ biomass, sex ated with digital photographs and measurements ratio, maturity at age, realised fecundity and gamete were taken with ImageJ (Schneider et al. 2012). Mor- viability. Some of these reproductive attributes are phometric characters include spore length, width directly related to fish condition and bioenergetic (W1, W2) and thickness (T1, T2), and length of the indices, which depend on diverse variables. Differ- large, intermediate and small polar capsules (PCs), ences in fish condition are often evaluated using rel- measured according to Giulietti et al. (2019). Ratios ative liver weight and body condition, since these are between spore width 1 and width 2 (W1:W2), thick- related to fecundity and maturation and are easy to ness 1 and thickness 2 (T1:T2), length of the large PC quantify (Rideout et al. 2006). A decrease in fish con- and length of the intermediate PC (LPCL:IPCL), as dition is linked to reduced fecundity and skipped well as length of the large PC and the length of the spawning (Rideout & Rose 2006, Rodgveller 2019). small PC (LPCL:SPCL) were calculated. The present study reports the infection of large vitel- logenic ovaries in apparently healthy wild fish by the myxozoan Kudoa sp., causing a strong decrease in 3. RESULTS reproductive potential without a noticeable decrease in body condition. A total of 361 (219 females and 142 males) Trachu- rus picturatus gonads were histologically analysed. Twenty-one females, ranging from 25.2 to 45.1 cm, 2. MATERIALS AND METHODS showed advanced vitellogenic oocytes infected with Kudoa sp., which appeared as an intense dark blue From September 2018 to August 2019, monthly in the Giemsa-stained sections (Fig. 1). Infected fe - samples of blue jack mackerel were acquired from males were detected mainly during and at the end of commercial vessels operating on the west coast of the reproduction season, from February to August, Portugal. Samples were obtained from fresh fish, and and for some months represented over 25% of the total length (TL, to the nearest 0.1 cm), eviscerated reproductive females analysed (Table 1). Only fe - weight (EW, to the nearest 0.01 g), gonad and liver male gonads were parasitized with Kudoa sp. weights (GW and LW, respectively, to the nearest No differences in the gonad macroscopic features 0.01 g), and sex were recorded. were evident between infected and uninfected Gonads were fixed in 10% buffered formalin for spawning-capable females (Fig. 2), and significant posterior histological analysis. For this, a portion of differences among indices were only noticed for each gonad was dehydrated with ethanol, embedded the GSI (GSI: FModel = 2.9950, p = 0.0090; HSI: in methacrylate, sectioned at 3 µm and stained with FModel = 0.7117, p = 0.6298; K: FModel = 0.0927; p = toluidine blue. Images were captured with a Leica 0.9780), with higher mean values found for infected DFC 290 digital camera attached to a Leica DM 2000 females in some months (Fig. 3). light microscope. Gonads were histologically exam- Kudoa sp. spores were stellate in shape with 4 ined for maturity phase assignment according to the unequal pyriform PCs in apical view (Fig. 1). The standardised terminology proposed by Brown-Peter- spore and PC measurements of the present species son et al. (2011). During histological analyses, the (mean, SD and range) are given in Table 2. presence of possible Kudoa sp. myxosporean para- Kudoa sp. spores appeared at the periphery of the sites was detected on advanced vitellogenic oocytes. oocytes, near the zona radiata (Fig. 4a), developing To confirm the presence of the parasite, subsamples tubular ramified plasmodia from the inner mem- of the same gonads were embedded in paraffin wax brane toward the nucleus (Fig. 4b). Mass infection and stained with Giemsa. of oocytes with only primary growth oocytes remain- The gonadosomatic index (GSI = 100 × GW / EW), ing in the ovary (Fig. 4c) was observed in several hepatosomatic index (HSI = 100 × LW / EW) and Ful- specimens. Neves et al.: Kudoa infection of Trachurus picturatus ovaries 49

Fig. 1. (a) Giemsa-stained histological section of infected Trachurus picturatus oocyte with (b) a detailed aspect of Kudoa spores. Scale bars = (a) 100, (b) 10 µm

Table 1. Monthly occurrence of Trachurus picturatus females infected with Kudoa sp., spawning-capable (SC) females and total females from the western Portuguese coast

Sep Oct Nov Dec Jan Feb Mar Apr May Jun Jul Aug

Kudoa-infected females 1 0 1 0 0 2 7 2 2 2 1 3 SC females 0 0 0 0 17 17 25 4 0 3 1 0 Total females 22 19 15 10 32 26 35 23 27 26 11 15

4. DISCUSSION ad vanced vitellogenic oocytes, which will lead to a reduction of the species SRP and to consequent Myxozoans of the genus Kudoa, with over 100 impacts on fisheries productivity. species described, infect marine and estuarine fish In the present study, no macroscopic evidence species worldwide (Giulietti et al. 2019). Most of of infection was noticed in fresh individuals, with these parasites are described to infect the skeletal an absence of post-mortem liquefaction of infected musculature, but some occur in the brain, heart, fish tissues and no visible difference between gills, kidney, ovary or intestines (Eiras et al. 2014). ovary tissue from infected and uninfected females, Some species are known to cause economic loss a common fact, as gonad infections do not gener- through the induction of post-mortem liquefaction ally produce obvious external signs (Sitjà-Bobadilla of fish muscles and by the production of macro- 2009). scopic cysts in the fish musculature reducing its The presence of Kudoa infection was only detected market value (Lom & Dyková 2006). The incidence in histological slides, after all gonad tissue had been of parasites in fish gonads is difficult to assess fixed in 10% buffered formalin. This prevented the since, as stressed by Sitjà-Bobadilla (2009), gonads assessment of fresh Kudoa sp. samples. A few studies are not always examined in parasite monitoring, have reported the occurrence of ovary infection thus restricting the frequency of myxosporeans (Swearer & Robertson 1999, Mansour et al. 2013, described for gonads despite their huge diversity 2015) and all described a new Kudoa species. Most and vast host number. In fact, Sitjà-Bobadilla (2009) Kudoa species are host specific, infecting fish of a mentions only 2 myxosporean species specifically particular family or even a single species (Whipps & affecting the ovaries of marine fish, although more Kent 2006) and many also seem to be tissue specific. species have since been described. Infection of the K. nova, K. quadratum and K. trachuri have been re - ovary, as described here, results in a total loss of ported to infect Trachurus spp. musculature (Moran 50 Dis Aquat Org 141: 47–52, 2020

Fig. 2. Macroscopic (top) and microscopic (bottom) aspect of (a) an infected and (b) an uninfected ovary of Trachurus picturatus

Fig. 3. Mean and standard deviation of the (a) gonadosomatic index (GSI), (b) hepatosomatic index (HSI) and (c) Fulton’s con- dition factor (K) for uninfected Trachurus picturatus females in the spawning-capable stage (black dots) and those with Kudoa sp. infected ovaries (open dots) Neves et al.: Kudoa infection of Trachurus picturatus ovaries 51

Table 2. Spore morphometry of Kudoa sp. found in ovaries of The ovary infection by Kudoa sp. detected in sev- Trachurus picturatus from the western Portuguese coast. eral T. picturatus specimens is most certainly affect- Measurements (mean, SD and range, given in µm) were ing the SRP on the Portuguese coast. Adlerstein & made on histological slides of formalin-fixed ovary tissue. L: spore length (lateral view); W1, W2: spore width (apical Dorn (1998) found a significant decrease, as much as view); T1, T2: spore thickness (apical view); LPCL: length of 90%, in the average fecundity of Merluccius produc- large polar capsule; IPCL: combined length of the 2 interme- tus infected by K. paniformis in the muscle. No quan- diate polar capsules; SPCL: length of the small polar capsule tification of fecundity decrease is available for gonad infection, although such an infection would be Mean SD Range expected to have a great impact on fish fecundity (Sitjà-Bobadilla 2009 and references therein). L 3.01 0.42 2.31−3.91 W1 4.9 0.55 3.31−5.31 One of the main assumptions in fishery assessments W2 4.19 0.63 3.24−5.86 for iteroparous fish species is that, after reaching T1 3.01 0.44 2.3−3.99 maturity, individuals reproduce annually (Macchi et T2 2.67 0.32 2.12−3.49 al. 2016). Nevertheless, the failure to spawn in con- LPCL 1.36 0.15 1.09−1.80 IPCL 1.21 0.11 1.01−1.56 secutive years (skipped spawning) has been observed SPCL 1.04 0.11 0.81−1.33 in many species (Rideout & Tomkiewicz 2011) and W1:W2 1.00 0.09 0.82−1.23 has already started to be incorporated in stock-recruit T1:T2 1.15 0.10 1.00−1.48 functions (e.g. Brooks 2013). Another suppressed re- LPCL:IPCL 1.13 0.13 0.83−1.57 LPCL:SPCL 1.31 0.17 1.03−1.78 production event is now described, which implies a decrease in the reproductive potential of the Por- tuguese population of T. picturatus. In this case, bio- et al. 1999, Campbell 2005, Matsukane et al. 2011), logical indices do not seem to act as a signal to the re- while K. azevedoi has been detected in the ovary of production failure, as the affected individuals can Trachurus species (Mansour et al. 2013). only be identified by histological examination of the The Kudoa spores found in the present study are of gonads. An overestimation of annual egg production small size, which appears to be characteristic of the could occur if such reproductive abnormalities are Kudoa species infecting ovaries (Mansour et al. 2013, not taken into account when models are applied. Fur- 2015). However, the fact that measurements in the ther studies are necessary in order to clearly quantify present study were made on fixed ovary tissue will reproductive losses and to assess its occurrence in certainly bias them negatively, hindering a substan- different populations of T. picturatus. tiated comparison of the measurements with those reported by other authors. K. azevedoi has been de - Acknowledgements. This study was partially supported by scribed as an ovarian parasite of a close species, T. the European Maritime and Fisheries Fund MAR2020 pro- trachurus (Mansour et al. 2013), and although it is ject ‘VALOREJET: Valorização de espécies rejeitadas e de possible that the same species infects T. picturatus, baixo valor comercial’, MAR-01.03.01-FEAMP-0003 and by Fundação para a Ciência e Tecnologia through research additional studies reporting fresh spore measure- contracts attributed to V.S. (CEECIND/02705/2017) and ments and rDNA sequence analyses will be needed A.R.V. (CEECIND/01528/2017), and strategic project UID/ to clarify the species definition. MAR/ 04292/2019.

Fig. 4. Toluidine blue-stained histological sections of Trachurus picturatus ovaries infected with Kudoa sp. showing infected (IO) and uninfected oocytes (NO). (a) Kudoa spores condensed at the periphery of an advanced vitellogenic oocyte; (b) spore proliferation with yolk fragmentation; (c) massive infection with complete degradation of the oocyte. PG: primary growth oocyte; scale bars = 200 µm 52 Dis Aquat Org 141: 47–52, 2020

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Editorial responsibility: Stephen Feist, Submitted: April 30, 2020; Accepted: July 16, 2020 Weymouth, UK Proofs received from author(s): September 11, 2020