The role of thyroid-stimulating hormone (TSH) in the regulation of peripheral thyroid function in non-mammalian species is still poorly understood. Thyroxine (T₄), the principal hormone released from the thyroid gland in response to TSH stimulation, circulates with a robust daily rhythm in the sciaenid fish, red drum. Previous research has suggested that the red drum T₄ cycle is circadian in nature, driven by TSH secretion in the early photophase and inhibited by T₄ feedback in the early scotophase. To determine whether TSH is produced in a pattern consistent with driving this T₄ cycle, I developed quantitative real time RT-PCR (qPCR) techniques to quantify the daily cycle of expression of the pituitary TSH subunits GSU[alpha], and TSH[beta]. I found that pituitary TSH expression cycled inversely to, and 6-12 hours out of phase with, the T₄ cycle, consistent with the hypothesis that TSH secretion drives the T₄ cycle. To examine the potential role of deiodinases in negative feedback regulation of this TSH cycle, I also utilized qPCR to assess the pituitary expression patterns of the TH activating enzyme outer-ring deiodinase (Dio2) and the TH deactivating enzyme inner ring deiodinase (Dio3). Whereas Dio2 was not expressed with an obvious daily cycle, Dio3 was expressed in the pituitary mirroring the TSH cycle. These results are consistent with T₄ negative feedback on TSH and suggest that TH inactivation by pituitary cells is an important component of the negative feedback system. To further examine the TH regulation of this Dio3 cycle, I developed an immersion technique to administer physiological doses of T₃ and T₄ in vivo. Both hormones persist in static tank water for at least 40 hours. Immersion in 200ng/ml T₄ significantly increased both plasma T₄ and T₃ within physiological ranges above control at 4.5 hours. Immersion in 100ng/ml T₃ increased plasma T₃ within physiological ranges over control by 22 hours while significantly decreasing plasma T₄ below control, presumably through inhibition of TSH secretion. T₄ also significantly inhibited the expression of the TSH [alpha] and [beta] subunits at 4.5 and 22 hours of immersion whereas T₃ immersion significantly inhibited the expression of the [alpha] and [beta] subunits of TSH by 22 hours. Both Dio2 and Dio3 expression were significantly diminished by T3 and T₄ at 22 hours. Inhibition of circulating THs with the goitrogen methimazole significantly increased the expression of TSH. These results indicate that both T₄ and T₃ are capable of negative feedback regulation of TSH expression in red drum on a time scale consistent with the T₄ daily cycle, and further support Dio3 destruction of THs in the pituitary, potentially regulated by circulating T₄, as a critical component of negative feedback on TSH. This study supports the importance of central mechanisms acting through pituitary TSH secretion in regulating thyroid function in red drum.

This collection of studies was designed to understand the mechanisms and consequences of competition among early life stages of red drum (Sciaenops ocellatus), with a special attention to asymmetric competition and multiple-breeding strategy of parents. The overall hypotheses were that (1) red drum larvae show behavioral interactions, and the magnitude of these interactions is explained by the sizes of the competing individuals, (2) red drum larvae compete for food and habitat use, and the competition is asymmetric as determined by size and behavioral interactions, and (3) the parents can reduce negative effects of competition among larvae and increase larval survival by altering their multiple-spawning traits. The laboratory experiments showed the existence of sociality in red drum larvae. The magnitudes of aggressive behavior, vigilant behavior and shoaling behavior were explained by a combination of absolute and relative phenotypes of competing individuals, such as body lengths and body condition. A new method of combining the laboratory experiments and the foraging theory disentangled interference competition for food from exploitative competition, revealing that both absolute and relative body lengths of competing individuals influence feeding efficiencies. In addition, vigilant behavior decreased feeding efficiency regardless of body sizes. The competitor size and behavioral interactions between individuals had different effects on competition for habitat use: the existence of larger individuals prevented the newly settled larvae from entering preferred habitats, whereas the aggressive behavior from the occupants of preferred habitats moved newly settled larvae to the unpreferred habitats. Finally, computer simulations using an individual-based model revealed that as interference competition among larvae increased, the number of spawning events and the spawning interval of the parents increased to lessen competition and maximize total survival of offspring. / text

Survival of teleost larvae requires growth and development which depend upon endocrine processes. In this dissertation I have examined the ontogeny of elements of the thyroid hormone (TH) and corticosteroid (CS) systems in red drum (Sciaenops ocellatus). Basal cortisol production was first detected 3 days post-hatch (DPH) and a cortisol stress response was present from 6 DPH forward. Changes in steroidogenic enzyme mRNA (CYP11B and CYP21) levels did not correlate with these events. The time necessary to reach peak cortisol levels as well as return to basal levels declined as larvae developed. A second set of studies examined ontogenetic patterns in levels of mRNAs encoding thyroid (soTR[alpha] and soTR[beta]) and corticosteroid (soGR) hormone receptors and assessed the regulation of these mRNAs by exogenous triiodothyronine (T3) and cortisol. soTR[alpha], soTR[beta] and soGR were expressed in all stages of red drum examined. soTR[alpha] levels increased during the time when surging TH levels have been reported. soTR[beta] levels did not differ significantly during development. soGR levels were strongly correlated with those of soTR[alpha]. T3 up-regulated soTR[alpha] and soTR[beta] levels in 7 DPH, but not older larvae. Cross-regulation of receptor mRNAs by exogenous treatment with T3 or cortisol was not observed. Finally, I assessed the influence of a common herbicide, atrazine, on receptor mRNA, TH levels and growth of red drum. In two experiments, red drum exposed to environmentally relevant levels of atrazine did not alter hormone receptor mRNA levels, or TH content. However, atrazine did depress growth in some instances. In conclusion, the expression patterns of hormone receptor mRNA in embryos suggests receptor proteins could be activated by maternal hormones prior to the onset of endogenous hormone production. A correlation between soTR[alpha] and soGR mRNA levels suggests coordinated function of TH and CS systems, although regulatory interactions between these systems were not evident under the conditions in this study. Patterns in soTR[alpha] and soTR[beta] mRNA levels support an important role for TH in the larval to juvenile transformation of red drum larvae. The results also support growing evidence indicating atrazine exposure effects larval growth and may impact their survival in the wild. / text

Hypervariable, nuclear-encoded microsatellites were used to (i) estimate genetic effective size (Ne) of red drum spawning over a two-week period in nine brood tanks at a TPWD hatchery; (ii) estimate heritability of early-larval growth and of growth rate and cold tolerance of juveniles; and (iii) test Mendelian segregation and independent assortment of 31 nuclear-encoded microsatellites. Assuming all tanks contributed equally to an offspring population, the maximum (expected) and observed Ne over the nine brood tanks was 43.2 and 27.0, respectively. The estimate of Ne based on observed variation in family size was 19.4. Simulations indicated that over a limited time period the simplest approach to maximizing Ne for a release population would be to utilize equal numbers of progeny from each brood tank. A family (genetic) effect was found to contribute significantly to the variance in early larval growth, juvenile growth rate, and cold tolerance. Estimates of narrow-sense heritability for these three traits were 0.07 +- 0.03, 0.52 +- 0.21 and 0.20 +- 0.10 (two growth intervals measured), and 0.30 +- 0.11, respectively, under the genetic models employed. The relatively low estimate of heritability for early larval growth suggests that genetic improvement for this trait likely would be slow. The heritability estimates for juvenile growth rate and cold tolerance, alternatively, suggest that genetic selection for these traits could be effective. Segregation at all 31 microsatellites fit Mendelian expectations for autosomal loci; a null allele was inferred at two of the microsatellites. Results from pairwise tests of independent assortment demonstrated that 20 of the 31 microsatellites could be placed into seven linkage groups. Additional linkage groups inferred from a prior study increased the number of inferred linkage groups in red drum to nine, with a range of two - five (avg. = 2.78) microsatellites in each linkage group. The remaining 11 microsatellites tested in this study assorted independently from all other microsatellites, suggesting the possibility of 11 additional linkage groups.

As aquaculture continues to expand, protein sources have become more costly and less available. Simultaneously, lipid-extracted algal meals (LEA) (co-products of bio-diesel production) are becoming increasingly available as alternative sources of energy are investigated. By integrating LEA into aquaculture diets, feed prices could be lowered and bio-diesel production would have an additional revenue stream. Three feeding trials each of 7 weeks duration were conducted to evaluate five different algal meals as partial replacements for fishmeal and soy protein concentrate in diets for juvenile red drum (Sciaenops ocellatus). Apparent digestibility coefficients (ADCs) for CP and energy in various LEAs also were determined with red drum and hybrid striped bass (HSB). In the first trial, whole algae meal and LEA derived from Navicula sp. replaced 5 or 10% of the crude protein (CP) in the reference diet. Weight gain, feed efficiency (FE), hepatosomatic index (HSI), as well as protein and energy retention were not significantly (P>0.05) affected by the dietary treatments. Algal inclusion significantly affected the ADCs of the various dietary treatments for dry matter, CP, and energy. The inclusion of ash in the form of diatomaceous earth improved digestibility of protein as well as weight gain, survival and FE. A second feeding trial evaluated LEA derived from Chlorella sp. processed at high temperatures, replacing 5, 10, 20, and 25% of the CP in the reference diet. Weight gain, FE, survival, and protein efficiency ratio (PER) were significantly reduced at substitution levels of 20 and 25%. The third feeding trial evaluated LEA derived from Nanochloropsis salina, replacing 5, 7.5, 10, and 15% of the CP in the reference diet. Weight gain, FE, survival, and PER were significantly affected by some dietary treatments, with the 15% substitution levels causing significant reductions in performance. Intraperitoneal fat ratio and whole-body lipids were significantly lower in the fish fed the 15% substitution diet. Based on the results of these experiments, replacement of up to 10% of CP from fishmeal and soy protein concentrate with LEA was possible without causing substantial reductions in fish performance, and the whole algae product provided a more nutritious product. Red drum and HSB showed similar responses in their ability to digest CP and energy from the various algal products, although ADCs varied greatly among the different products and processing methods.

Results from the present study found for the first time that the use of bacterial strains of Lactobacillus acidophilus, Lactobacillus casei, Bifidobacterium thermophilum, and Enterococcus faecium from the commercial product PrimaLac® had the potential to act as a possible probiotic for juvenile red drum. The addition of PrimaLac® probiotics [whether as a water-soluble probiotic (WSP) or in a probiotic enhanced starter feed (PESF)] reduced mortality (%M), enhanced growth rates (MW, LT, SGR, and DGR), improved feed conversion efficiencies (FCEs), and physical condition factors in the juvenile red drum. Improvement was quantified using external morphological condition indices (MCIs) and blood physiological condition indices (PCIs). Results showed a strong positive relationship between MCIs and PCIs, which suggested that probiotics treated fish were in better health (lower MCIs) with lower fasting blood glucose and lactate levels than control fish. Addition of probiotics also resulted in improved water quality (lower nitrate, nitrite, and ammonia) in the treatment tanks compared to controls. Application of PrimaLac® probiotics on the morphology and histology of three different regions of the intestine (proximal, mid-and distal) improved intestinal length (Li), mass (Mi), and digesta mass (Md). In 5μm histological sections examined for differences among treatments (probiotics vs. controls), five variables within each intestinal region were also quantified: (i) perimeter ratio (PR) between the internal perimeter (IP) of the intestinal lumen and the external perimeter (EP) of the intestine where, PR = IP/EP; (ii) width of lamina propria (wLP); (iii) thickness of the muscle layer (tM); (iv) the number of goblet cells per segments (GC); and (v) height of enterocytes (hE). Overall, Li and Mi significantly increased by 13.4 ± 1.2% and 11.8 ± 0.9%, respectively, and Md decreased by 33.2 ± 1.21% compared to controls. Significant increases in microvilli heights, PR (by 21.3 ± 1.2%), tM (by 18.4 ± 0.9%), and increased numbers of GC (by 35.1 ± 2.8%) provided evidence that bacteria species from PrimaLac® (whether in WSP or PESF) affected the intestinal histomorphology of juvenile red drum. Collectively, these changes were partially responsible for increased feed assimilation (supported by reduced digesta) and improved overall juvenile red drum growth and condition. These results will add significantly to the limited database on how probiotics, as possible standard practice at the fish farm, may offer significant benefits for fish production and fish health. Addition of probiotics will also be of benefit to most aquaculture species and not only for red drum, but also for other high-value carnivorous species, such as Atlantic salmon, without threat to natural populations or damaging fisheries-based ecosystems that are already on the edge of collapse from over fishing and pollution.

The main objective of this thesis was to gain new insights in three of Florida’s high value food and stock enhancement finfish nutrition (Common snook, Florida pompano and red drum) to improve larviculture protocols. The main bottleneck in snook production is the extremely low larval survival rate, which hinders subsequent research. This work first focused on the source of the larvae by looking at potential nutritional deficiencies in captive broodstock. The lipid composition of wild and captive common snook broodstock were compared to identify disparities and gain the information necessary for the formulation of a suitable diet for captive stocks. Results showed that captive snook lipid content was significantly higher than that of wild fish. However, cholesterol and arachidonic acid (ARA) levels were significantly lower compared to wild broodstock, with potential impact on steroid and prostaglandin production, reproductive behavior and gametogenesis. Eggs from captive broodstock incorporated high docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) levels and low ARA levels. Consequently, ARA/EPA ratio in captive eggs was more than half of that in wild eggs (2.3 ± 0.6 and 0.9 ± 0.1 respectively), with a probable perturbation in eicosanoid production and adverse effects on embryo and larval development. The large differences observed between wild and captive broodstock most likely contributed to the reproductive dysfunctions observed in captive snook broodstock (e.g. incomplete oocyte maturation, low milt production and poor egg quality). In addition, the presence of hydrocarbons was detected in the liver of most of the wild snook sampled. This requires further investigation to identify the source of the contamination, monitor a potential impact on reproductive performances and protect the species habitat. Another major bottleneck in marine fish rearing occurs during the transition from endogenous feeding to exogenous feeding, with mass mortality events linked to inadequate first feeding diets. To gain insight on the early fatty acid requirements and mobilization of pompano and snook larvae, the pattern of conservation and loss of fatty acids from the yolk sac during the endogenous feeding period and subsequent starvation was studied. In both species, fatty acids were utilized as an energy source after hatching. Mono-unsaturated fatty acids were catabolized, while saturated and poly-unsaturated fatty acids were conserved. High levels of arachidonic acid (ARA) in pompano and snook eggs (respectively 2.44 ± 0.1 and 5.43 ± 0.3 % of total fatty acids), as well as selective retention in the unfed larvae, suggested a high dietary requirement for this fatty acid during the early stages of larval development. The effect of an ARA supplementation was therefore investigated in snook larvae at the rotifer feeding stage. Larvae receiving the supplementation did incorporate higher levels of ARA, and DHA/EPA and ARA/EPA ratios were successfully modified to match those observed in wild eggs. No significant improvements in growth or survival were observed, however the success in fatty acid profile modification suggest a possible impact of the supplementation on a longer period of time and a possible effect on stress resistance. Probiotics have been shown to enhance larval performances of several species and this strategy was therefore investigated to evaluate a potential impact on Florida pompano, red drum and common snook larvae. The effect of a commercial mix of Bacillus sp. was studied on larval survival, growth and digestive enzyme activities. Larvae were fed either live feed enriched with Algamac 3050 (Control), Algamac 3050 and probiotics (PB), or the previous diet combined with a daily addition of probiotics to the tank water (PB+). Microbiological analyses were performed at the end of the pompano trial. Numbers of presumptive Vibrio sp. were low and not statistically different between treatments, therefore no additional microbiological analyses were performed on the system. At the end of the pompano and snook trial, standard lengths of larvae from the PB and PB+ treatments were significantly greater than for the control larvae. For both pompano and snook, trypsin specific activity was higher in PB and PB+ larvae compared to the control larvae. Similarly, alkaline phosphatase activity was higher for the pompano larvae fed the PB and PB+ treatments and for the snook larvae fed the PB+ treatment compared to the control larvae. No enhancement of growth or digestive enzymes activities was observed in red drum larvae. Yet, no negative effects were noticed and a longer trial period and the study of additional parameters could reveal different effects. In all three species, survival was not affected by the supplementation; however, stress exposure should be further investigated as the supplementation may strengthen the larvae, especially pompano and snook larvae where the Bacillus sp. supplementation appears to promote growth through an early maturation of the digestive system. Another key challenge in marine fish larval rearing resides in weaning the larvae onto dry micro-diets. This step is commonly concurrent with larvae metamorphosis into juveniles, with extensive morphological and physiological changes that are likely to influence nutritional requirements. In the present project, three microdiets were tested on weaning of Florida pompano larvae: Otohime, Gemma and a reference diet LR803. The experimental system was stocked with 11-day-old larvae, which were co-fed micro-diets and live food from 11 dph to 17 dph then micro-diets only until 28 dph. Survival from 11 dph to 28 dph was similar for all treatments, with an average of 33 %. At the end of the trial, the Gemma larvae were significantly longer and heavier than larvae fed the other diets. Fatty acid composition of the diets and larvae varied significantly between treatments. The Gemma larvae incorporated the lowest amount of eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and arachidonic acid (ARA). However, they presented the highest DHA/EPA and ARA/EPA ratios, supporting the concept that the proportions of polyunsaturated fatty acids are of greater importance than their absolute amount. Results from the enzyme analysis showed that fishmeal is a suitable main source of protein for Florida pompano larvae and demonstrated the full functionality of the pancreas at 16 days post hatch. These results provide the basis of a suitable weaning diet for pompano larvae and indicate the possibility of a weaning time prior to 16 days post hatch, which is of high interest in commercial production. Overall, this research provides new data on common snook, pompano and red drum nutritional requirements with results that can be directly applied to help overcome major bottlenecks in the hatchery phase and improve rearing protocols.

Bayou St. John (BSJ) and City Park Lakes and Lagoons (CPLL) are urban waterways in New Orleans, Louisiana. I studied habitat selection of red drum in BSJ, and fish assemblage change in BSJ and CPLL over 40 years. Temperature was found to be the best predictor of red drum habitat selection in Bayou St. John, while salinity and change in depth also were found to be good predictors for certain sites. Potential prey item abundance did not appear to influence habitat selection. Using data from 1971 – 2010, nearshore habitats in CPLL were affected by Hurricane Katrina, but have sense recovered and nearshore habitats in BSJ were found to have decreased diversity. Pelagic habitats in both areas were found stable across 40 years. Since 2006, nearshore assemblages were similar for CPLL and BSJ with a decrease in fishes from Order: Cyprinodontiformes and an increase in other fishes seen across years.

Quantifying the impacts of restoration on coastal waterways is crucial to understanding their effectiveness. Here, I look at the impacts of multiple restoration projects on urban waterways within the city limits of New Orleans, LA, with an emphasis on the response of fishes. First I report the effects of two projects designed to improve exchange down estuary on the hydrologic characteristics of Bayou St. John (BSJ). Within BSJ, flow is dominated by subtidal wind driven processes. Removal of an outdated flood control structure did not appear to alter exchange in BSJ, but removal combined with sector gate openings did. I also refined a three dimensional hydrodynamic model of this system to have accurate predictions of velocity and elevation. Temperature and salinity were difficult to constrain with this model. Solutions of this model were used to compare flow metrics, along with linearly interpolated temperature, and other variables to Red Drum (Sciaenops ocellatus) activity and movement patterns. Relationships between Red Drum activity and velocity suggested a response to subtidal, wind driven flow. Overall, high Sedentariness, a measure of inactivity, was found suggesting high levels of site fidelity. Higher mean Sedentariness during the night was also found. I also used a pseudo-BACI design to analyze the fish assemblage response to removal of an outdated flood control structure and the impacts of sector gate openings on fish guild species richness in BSJ. Limited differences were found when comparing fish assemblages before and after removal, but these differences were likely due to a decrease in salinity not restoration efforts. No significant differences in Freshwater or Estuarine fish guild species richness was observed for any of the control or impact sites. Marine fish species richness was found to be higher immediately following sector gate openings at the site closest to the structure, suggesting an initial pulse of young marine organisms is provided via these events. The findings here can be used to optimize management of exchange flow in coastal impounded waterways.

Le poisson est un produit très périssable dont l’altération résulte essentiellement de la croissance bactérienne. Comparé aux régions tempérées, peu d’études portent sur le microbiote d’altération des poissons tropicaux. En Martinique, le thon jaune (Thunnus albacares) et l’ombrine ocellée (Sciaenops ocellatus) représentent des poissons d’intérêt pour les filières pêche et aquaculture. Dans le but de mieux connaître le microbiote d’altération de ces poissons, des analyses culturales et aculturales (séquençage de nouvelle génération des amplicons d’ARNr 16S, Illumina MiSeq) ont été réalisées.Une grande diversité d’espèces bactériennes a été retrouvée dans le thon et l’ombrine fraîchement pêchés (104 et 887 OTUs, respectivement) et la plupart d’entre elles sont communément isolées des poissons (Chryseobacterium, Burkholderia, Flavobacterium, Psychrobacter, Arthrobacter, Staphylococcus). Certaines, comme Ralstonia sp. et Rhodanobacter terrae, en quantité importante dans le thon frais, sont plus atypiques. Au cours de l’entreposage du thon sous-glace, Pseudomonas et Brochothrix deviennent dominants. L’emballage sous atmosphère modifiée (MAP) ou sous vide (VP) entraine clairement la sélection de Brochothrix dans un cas et d’un mélange de Brochothrix, bactéries lactiques (Lactococcus piscium, Carnobacterium maltaromaticum) et d’entérobactéries (Hafnia paralvei) dans l’autre, et ne permet pas une augmentation significative de la durée de conservation. Pour les filets d’ombrine, peu de différences sont observées entre MAP et VP dont le microbiote se compose essentiellement de bactéries lactiques (Carnobacterium spp., Vagococcus spp., Lactococcus spp., Leuconostoc spp.). La durée de conservation est étendue de 15 jours par rapport au poisson entier sous air.L’inoculation de différentes espèces bactériennes dans de la chair pauci-microbienne de thon ou d’ombrine a montré que Hafnia paralvei et Serratia spp. sont les espèces les plus altérantes. Brochothrix thermosphacta et Carnobacterium spp. produisent aussi des odeurs indésirables mais de façon plus modérée. Chez Pseudomonas, les espèces ne sont pas toutes altérantes et présentent même parfois des capacités à empêcher le développement des mauvaises odeurs induites par d’autres bactéries (Pseudomonas psychrophila/fragi) et à dégrader l’histamine (Pseudomonas cedrina, Pseudomonas plecoglossicida/monteilii). En parallèle, des tests sensoriels et des dosages physico-chimiques ont également été réalisés pour comprendre les conséquences de la croissance bactérienne et identifier des indicateurs fiables pour l’évaluation du degré d’altération des produits. / Fish is a highly perishable product and spoilage is mainly due to the bacterial growth. Compared to temperate regions, few studies examined the spoilage microbiota of tropical fish. In Martinique, yellowfin tuna (Thunnus albacares) and red drum (Sciaenops ocellatus) are essential fish of fisheries and aquaculture sectors. For a better characterization of the microbial ecosystem, culture-dependent and culture-independent (next-generation sequencing of 16S rRNA amplicons, Illumina MiSeq) methods were carried out.A wide diversity of species was found in freshly caught tuna and red drum (104 and 887 OTUs, respectively) and most of them are commonly isolated from fish (Chryseobacterium, Burkholderia, Flavobacterium, Psychrobacter, Arthrobacter, Staphylococcus). Others, such as Ralstonia sp. and Rhodanobacter terrae, largely present in fresh tuna, are less familiar. During the ice-storage of tuna, Pseudomonas and Brochothrix became dominant. The modified atmosphere packaging (MAP) and vacuum packaging (VP) clearly leaded to the selection of Brochothrix in one case and to a mixture of Brochothrix, lactic acid bacteria (Lactococcus piscium, Carnobacterium maltaromaticum) and enterobacteria (Hafnia paralvei) in the other case, and not conduct to a significant increase of the shelf-life. For red drum fillets, few differences were observed between MAP and VP with a microbiota essentially composed by lactic acid bacteria (Carnobacterium spp., Vagococcus spp., Lactococcus spp., Leuconostoc spp.). The shelf-life was extended by 15 days compared to the whole fish ice-stored.The inoculation of different bacterial species into the pauci-microbial flesh of tuna or red drum showed that Hafnia paralvei and Serratia spp. were the most spoiling bacteria. Brochothrix thermosphacta and Carnobacterium spp. produced more moderate undesirable odors. Among the Pseudomonas genus, not all species induced spoiling effects and some of them are even able to prevent the development of unpleasant odors from other bacteria (Pseudomonas psychrophila/fragi) and to degrade histamine (Pseudomonas cedrina, Pseudomonas plecoglossicida/monteilii).At the same time, sensory tests and physico-chemical assays were performed to understand the consequences of the bacterial growth and to identify reliable indices for the evaluation of the spoilage degree of the products.