High-energy diets for white sturgeon, Acipenser transmontanus Richardson

Four diets formulated for salmon were fed to 0.11 kg white sturgeon, Acipenser transmontanus Richardson, for 8 weeks. Dietary compositions ranged from 258 to 402 g lipid kg⁻¹, 535–378 g protein kg⁻¹ and 22.7–14.4 g protein MJ⁻¹ gross energy.
Fish in all treatments grew rapidly, utilized the diets efficiently and had body compositions similar to what has been found in previous studies, but there were some dietary effects. Sturgeon fed the diet with the highest lipid content and lowest protein/energy ratio had lower ( P < 0.05) specific growth rate, feed efficiency, and liver moisture and protein contents, and 6-phosphogluconate dehydrogenase activity, but higher liver lipid contents than fish fed the other three diets. Condition factor, organ to body weight ratios, whole-body and plasma concentrations of protein, glucose and triglyceride, and liver glucose-6-phosphate dehydrogenase, isocitrate dehydrogenase and malic enzyme activities did not differ significantly among dietary treatments. This suggests that white sturgeon subyearlings can utilize diets with high lipid contents (258–357 g kg⁻¹) to display good growth without major adverse effects on body composition and liver lipogenic enzyme activities.     

Outbreaks of Streptococcosis associated with Streptococcus iniae in Siberian sturgeon (Acipenser baerii) in China

Streptococcus iniae has emerged as an important fish pathogen over the past few decades causing high losses in aquaculture farms all over the world. At least 27 species of fish have been documented to be infected by S. iniae, including cultured and wild populations. In August and October 2013, a serious infectious disease characterized by body ulcer, internal organs haemorrhages and nodules showing on epicardium occurred on the Acipenser baerii farms in Ya'an country, China. Histological examination revealed a multisystemic, necrotising inflammatory response that was particularly marked in liver, kidney, heart and brain. Mass mortality (>40%) was observed in infected fish and two Gram‐positive cocci (Ab130920 and Ab131025) were obtained from kidneys and livers of diseased fish. Experimental infections with these two isolates resulted in marked symptoms in the sturgeons similar to those observed in natural outbreaks, and the LD₅₀ values of the two isolates were 5.1 × 10⁵ and 6.4 × 10⁵ cfu per fish respectively. The two microorganisms were identified as S. iniae through physiological and biochemical tests, 16S rRNA and lctO gene sequence analysis. Both two isolates showed a similar antibiotic susceptibility, which were sensitive to ampicillin, amoxicillin, cefazolin, amikacin, deoxycycline, florfenicol, azithromycin, ciprofloxacin, vancomycin and resistant to streptomycin, gentamicin, kanamycin, norfloxacin and sinomin (SMZ/TMP). Multiplex PCR assay for virulence genes showed both isolates possessed six main virulence genes: simA, scpI, pdi, pgm, cpsD and sagA genes. These results indicated that S. iniae could act as a pathogen of farmed A. baerii. This is the first report of S. iniae infection associated with mass mortality in A. baerii.     

Evaluation of lethal and non-lethal sampling methods for the detection of white sturgeon iridovirus infection in white sturgeon, Acipenser transmontanus (Richardson)

Pectoral fin tissue of white sturgeon was investigated as a potential non-lethal sample source for the detection of white sturgeon iridovirus (WSIV) infection. Histopathology and polymerase chain reaction (PCR) results using fin tissue were compared with the standard lethal histopathology sampling method that utilizes head tissue. Tissues for each of the three sampling methods were collected weekly for 8 weeks from individual sturgeon undergoing an experimental cohabitation challenge with fish infected with the Abernathy isolate of WSIV. Non-lethal fin histopathological evaluation did not reveal infection during the first 3 weeks of sampling, while non-lethal PCR and the lethal method were variable. However, all three sampling methods were equally capable of identifying infection from 4 to 8 weeks post-exposure. Of the survivors tested, all were negative by PCR and the lethal method, and only one fish was identified as being positive by non-lethal fin histopathology. In another experiment, all three sampling methods were applied to asymptomatic WSIV carriers in a case study conducted at the Kootenai Tribal Sturgeon Conservation Hatchery. Results showed that both lethal and non-lethal fin histopathology were equally effective in detecting infection, but PCR was unable to identify this strain of WSIV. Depending on the virus isolate, these results suggest that non-lethal sampling of fin tissue (histopathology or PCR) is comparable with the lethal sampling method at identifying WSIV infection once infection is established, and under certain circumstances may provide an alternative to lethal sampling.     

Dopaminergic influence on gonadotropin secretion in white sturgeon (Acipenser transmontanus)

The effects of dopamine on the secretion of two sturgeon gonadotropins (stGTH I and stGTH II) in sexually mature male white sturgeon (Acipenser transmontanus) were evaluated. In Experiment I, sturgeon were given intraperitoneal injections of physiological saline (PS), dopamine (100 mg kg⁻¹), the gonadotropin releasing hormone analog D-Ala⁶-des-Gly¹⁰-GnRH ethylamide (GnRHA) (10 μg/kg⁻¹), and a combination of GnRHa and dopamine. Fish receiving only GnRHa had significantly higher (p < 0.05) concentrations of plasma stGTH I and stGTH II compared to fish receiving PS, dopamine, or a combination of GnRHa and dopamine. Two hours following its administration, dopamine was effective in decreasing plasma concentrations of both stGTHs that were previously elevated by GnRHa. Dopamine or PS administered by themselves did not alter plasma concentrations of either stGTH. In Experiment II, sturgeon injected intraperitoneally with a combination of GnRHa and pimozide had significantly higher (p < 0.05) concentrations of plasma stGTH I and stGTH II compared to males receiving GnRHa or pimozide alone. While this effect of GnRHAa + pimozide was observed in the spring, no such potentiation was seen in these fish during the summer (Experiment III). These results represent the first evidence of dopaminergic inhibition of GnRH-induced pituitary gonadotropin secretion in Chondrostean fish.     

Estimating the ideal dietary indispensable amino acid pattern for growth of white sturgeon, Acipenser transmontanus (Richardson)

Juvenile white sturgeon (Acipenser transmontanus, Richardson) were fed or starved for 4 weeks, and their whole body amino acid composition determined at the beginning and the end of the study. The amino acid compositions of muscle, liver, gill and gastrointestinal tract, and the free amino acids in plasma of fed and starved fish were also determined. Very little variation was observed in the amino acid compositions of all the tissues examined after starvation. However, the plasma free amino acids were significantly (P < 0.05) decreased following starvation. On the basis of the amino acid composition and nitrogen retention values of the whole fish, daily indispensable amino acid requirements for protein accretion and obligatory nitrogen losses (mg amino acid 100 g body weighr^?1 day^?1) were estimated. The pattern of indispensable amino acids required for each of these is different, with the pattern for protein accretion containing much higher levels of lysine, but lower levels of tryptophan and sulfur amino acids. Individual amino acid retention rates varied from 33.3% of dietary tryptophan to 80.2% for arginine. The estimated dietary indispensable amino acid pattern for young growing white sturgeon (expressed as a percentage of total indispensable amino acids plus cystine and tyrosine) is: arginine, 14.0; histidine, 6.6; isoleucine, 8.8; leucine, 12.5; lysine, 15.8; methionine (plus cystine), 6.6; phenylalanine (plus tyrosine), 15.5; threonine, 9.7; tryptophan, 0.8; and valine, 9.7.     

First report of Streptococcus iniae in red porgy (Pagrus pagrus, L.)

This work describes the first isolation of Streptococcus iniae in red porgy, Pagrus pagrus (L.), and the first European isolation of this pathogen in gilthead seabream, Sparus aurata (L.). In both farmed fish species, infection resulted in lethargy, anorexia, abnormal swimming, exophthalmia and sudden death, with mortality rates of over 25% in red porgy and 10% in gilthead seabream. Beta-haemolytic Gram-positive cocci, catalase negative and oxidase negative, were isolated in pure culture from internal organs. Conventional and rapid identification systems, and 16S rRNA gene partial sequencing were used to identify the causative agent of the natural disease. LD50 trials were carried out to show the virulence of this isolated strain in these species, with values of 1.7 × 104 CFU per fish in red porgy and 1.32 × 105 CFU per fish in gilthead seabream. The most prominent lesions were meningoencephalitis and multifocal infiltration of macrophage cells in the kidney and spleen.     

Effects of steroid hormones on in vitro oocyte maturation in white sturgeon (Acipenser transmontanus)

The in vitro effects of several steroids on the maturation of intact white sturgeon (Acipenser transmontanus) ovarian follicles were investigated. At the highest concentration (1024 ng ml^–1 for the C21 steroids and 1139 ng ml^–1 for the C19 steroids), all of the C21 steroids tested, progesterone (P4), 17-hydroxyprogesterone (17OHP), 17,20-dihydroxy-4-pregnen-3-one (17,20-P), 17,(20,21-trihydroxy-4-pregnen-3-one 20-S), 11-deoxycortisol (S) and cortisol (F), as well as testosterone (T) induced germinal vesicle breakdown (GVBD) at 14 and 22 h. At 6 h, only P4 and 17,20-P induced maturation at the highest concentration (1024 ng ml^–1). At 14 and 22 h, 11-deoxycortisol was the most potent steroid inducer of GVBD followed by P4, 17OHP, 17,20-P, and 20-S. The steroid 11-hydroxytestosterone (11OHT) was completely ineffective at all concentrations and exposure times. The C21 steroids induced oocyte maturation at concentrations ranging from 4 to 1024 ng ml^–1, whereas T induced GVBD at 225 to 1139 ng ml^–1. Calculation of the mean effective concentration that induced 50% GVBD (EC50) from the 22 h incubations revealed the following order of potencies: S > P4 > 17OHP > 17,20-P > 20-S >> F > T. These bioassay results, together with previous findings on the endogenous production of steroids by ovarian follicles from gonadotropin-primed females, indicate that more than one steroid has a biological role in the resumption of meiosis in sturgeon oocytes and provides empirical evidence for P4, 17OHP, S, 20-S, and 17,20-P as maturation-inducing steroids in white sturgeon.     

Effects of feeding rates on growth performances of white sturgeon (Acipenser transmontanus) fries

Four 1‐week growth trials were conducted to determine the effects of feeding rates on the growth performances of white sturgeon (Acipenser transmontanus) fries 6–9 weeks after initiation of feeding. Six feeding rates with four replications were used in each of the four trials, and the feeding rates were 3.0–8.0, 2.0–7.0, 1.0–6.0 and 1.0–6.0% body weight (BW) per day in 1% increment, respectively. Number of fries per replicate and their initial BW (means ± SEM) were 60, 45, 30 and 30 and 2.8 ± 0.1, 4.5 ± 0.4, 8.5 ± 0.7 and 10.0 ± 0.7 g, respectively. The fries were kept at 18–19 °C and fed a commercial salmonid feed (488 g kg^?1 protein and 123 g kg^?1 fat). Mortality was low and unrelated to feeding rates. Final body weights, body weight increases, specific growth rates and feed efficiency were significantly (P < 0.05) affected by the feeding rates. Body moisture and lipid contents were significantly affected by feeding rates except body moisture content in trial II. Body protein contents were not affected by feeding rates except in trial III. Broken‐line analysis on specific growth rates indicated that the optimum feeding rates were 6.5 ± 0.4, 4.8 ± 0.2, 4.2 ± 0.1 and 3.8 ± 0.2% body weight per day, respectively, for white sturgeon fries 6–9 weeks after initiation of feeding.     

Growth response and acquired resistance of Nile tilapia, Oreochromis niloticus (L.) that survived Streptococcus iniae infection

This study determined the growth performance and acquired resistance of Nile tilapia, Oreochromis niloticus (L.) that survived Streptococcus iniae infection. Tilapia were challenged with three doses of S. iniae (8.8 × 10³, 8.8 × 10⁴ and 8.8 × 10⁵ CFU fish^?1 for low, medium and high challenges respectively). Groups of non‐injected and tryptic soy broth‐injected fish were maintained as controls. Significantly (P<0.05) higher mortality (45.0%) occurred in the high challenge treatment than in the low challenge treatment group (29.6%). The medium challenge group had mortality (36.3%) that did not differ significantly from the high or low treatment. Few fish died in the non‐injected and broth‐injected treatments (3.4% and 0.8% respectively). The tilapia that survived S. iniae infection used to assess growth performance were selected from survivors without gross clinical signs of disease. These fish were randomly stocked at a rate of 30 fish into each 57 L aquarium in triplicate and fed to apparent satiation for 8 weeks. No significant differences were detected in weight gain, feed intake, feed efficiency ratio or survival between S. iniae‐survived tilapia and the control treatments following the 8‐week growth performance trial. Following the 8‐week feeding study, tilapia were challenged with 1 × 10⁶ CFU fish^?1 of S. iniae to assess acquired immunity. Mean cumulative mortality was significantly higher (P<0.05) in the control treatments (41.7% for the non‐injected and 43.3% for the broth‐injected fish) than in the low, medium and high challenge treatments (7.4%, 3.3% and 8.3% respectively). Serum protein was significantly (P<0.05) elevated in the S. iniae‐survived tilapia that were subsequently challenged when compared with controls challenged for the first time. Agglutinating antibody titre was significantly higher in the fish in the medium and high challenge treatments, compared with the control fish challenged for the first time. The results suggest tilapia that survive S. iniae challenge without showing overt disease signs performed as well as non‐infected tilapia. Further, the S. iniae‐survived tilapia challenged following the 8‐week growth performance trial gained acquired resistance to homologous S. iniae challenge.     

Confirmation of ovarian homogeneity in post-vitellogenic cultured white sturgeon,Acipenser transmontanus

Assessing stage of oocyte maturity in female sturgeon by calculating oocyte polarization index (PI) is a necessary tool for both conservation propagation managers and caviar producers to know when to hormonally induce spawning. We tested the assumption that sampling ovarian follicles from one section of one ovary is sufficient for calculating an oocyte PI representative of oocyte maturity for an individual animal. Short-wavelength near-infrared spectroscopy (SW-NIR) scans were performed on three positions per ovary for five fish prior to caviar harvest. Samples of ovarian follicles were subsequently taken from the exact location of the SW-NIR scans for calculation of oocyte PI and follicle diameter. Oocyte PI was statistically different though not biologically relevant within an ovary and between ovaries in four of five fish. Follicle diameter was statistically different but not biologically relevant within an ovary in three of five fish. There were no differences in follicle diameter between ovaries. No statistical differences were observed between SW-NIR spectra collected at different locations within an ovary or between ovaries. These results emphasize the importance of utilizing both oocyte PI measurement and progesterone-induced oocyte maturation assays while deciding when to hormonally induce spawning in sturgeon females.     

豚链球菌和停乳链球菌类M蛋白及其抗原性分析

分别用热酸抽提法、碱抽提法和胃蛋白酶消化法分离豚链球菌NUF849的类M蛋白,通过SDS-PAGE和Western-blotting印迹法分析比较三种方法所提类M蛋白的组成和抗原性差异。结果表明,热酸抽提法分离制备的类M蛋白得率和纯度都较其它两种方法高,蛋白的抗原性保持较好。采用热酸提取法提取5株豚链球菌NUF633、NUF693、 NUF701、 NUF812、 NUF849和2株停乳链球菌SD、L2的类M蛋白,其SDS-PAGE图谱明显不同,种间差异明显, 豚链球菌主要类M蛋白的分子量分别为60、48、37、30、27、24和15kDa,停乳链球菌主要类M蛋白的分子量分别为68、48、42、37、29、26、23、21、19、17、15、14和11kDa。Western-blotting结果显示,兔抗豚链球菌血清可与豚链球菌的两条类M蛋白带结合,分子量分别为60和37kDa;与停乳链球菌的两条类M蛋白带结合,分子量为68和37 kDa。两种链球菌主要类M蛋白组成及其抗原性差异明显。     

Protection against heterologous Streptococcus iniae isolates using a modified bacterin vaccine in Nile tilapia,Oreochromis niloticus (L.)

Streptococcus iniae is a significant pathogen impacting aquaculture production worldwide. The objectives of this study were to determine whether a developed modified S. iniae (ARS-98-60) bacterin vaccine is efficacious in Nile tilapia, Oreochromis niloticus (L.), against challenge with heterologous isolates from diverse geographical locations and to evaluate protein and antigenic variability among the isolates tested. Two groups of tilapia (approximately 5 g) were intraperitoneally (IP) vaccinated with 100 μL of the vaccine or sham vaccinated with 100 μL of sterile tryptic soy broth and held for 28 days. Fish were challenged with each isolate by IP injection of 2–3 × 10⁷ CFU per fish using calcein to mark fish prior to cohabitation for challenge. The results demonstrated significant protection against all challenge isolates, and relative percent survivals ranged from 79% to 100%. SDS–PAGE analysis of whole-cell lysate proteins from the S. iniae isolates demonstrated similar protein profiles between 10 and 31 kDa and variation in profiles between 35 and 100 kDa. Western blot analysis using antiserum from vaccinated fish (ARS-98-60) demonstrated shared immunogenic proteins among all isolates in the molecular mass range of 22–35 kDa and high molecular mass material >150 kDa. The results suggest that the developed S. iniae vaccine has broad ranging protection among isolates exhibiting different protein profiles.     

Effects of feeding a high level of D-glucose on liver function in juvenile white sturgeon (Acipenser transmontanus)

Juvenile white sturgeon (Acipenser transmontanus) were fed three isonitrogenous and isoenergetic diets containing either 35% D-glucose (HC), a mixture of 20% dextrin and 10% cellulose (MC), or 23% cellulose (LC), to investigate the effects of dietary carbohydrate on liver function. After 8-week feeding, body weight gain of fish fed the HC diet was consistently higher than that of fish fed the MC and LC diets, but was not significantly different from the MC-fed fish. Fish fed the HC diet had significantly (p < 0.05) higher feed efficiencies and liver glycogen concentrations than fish fed the MC and LC diets. Sturgeon were injected intravenously with 10 mg kg^-1 body weight of sulfobromophthalein (BSP) and post-injection blood taken from the caudal vein at 15, 30, 60, and 120 min. No significant differences in plasma BSP concentrations were found among the treatments at these times. Plasma hemoglobin and activities of aspartate and alanine aminotransferase were not affected by the diets. This study suggests that the HC diet does not adversely affect liver function or weight gain. Inclusion of high dietary levels of digestible and inexpensive carbohydrates in commercial sturgeon feeds seems promising, but long-term feeding trials should be conducted to confirm this assertion.     

Effects of Spirulina and plant oil on the growth and lipid traits of white sturgeon (Acipenser transmontanus) fingerlings

The aim of this research was to evaluate the efficiency of diets with Spirulina and plant oils (POs) inclusion for white sturgeon weaning and their effects on the fatty acid (FA) composition of fish flesh. Three isoproteic (45%) and isoenergetic (21 MJ kg^?1 DM) diets were formulated: one fish meal‐based diet integrated with fish oil (FMO) and two 40% Spirulina meal‐based diets integrated with corn (SPC) or soybean (SPS) oils respectively. One hundred and thirty‐five white sturgeon fingerlings (mean weight 17.5 g) were stocked randomly in nine fibreglass tanks. At the end of the trial, which lasted 71 days, the growth performance traits and somatic indexes were determined. The chemical composition, gross energy and FA profile were determined on the fish fillets. No significant effects were observed for the growth performances or fillet chemical composition. The FA profile of the fillets reflected those of the diets. In particular, the fillets of the fish fed with the SPC and SPS diets were lower in n‐3 FA, due to the substitution of fish oil (FO) with POs. It is possible to replace FO and meal in sturgeon; therefore, Spirulina meal integrated with POs could be a good alternative to sturgeon diet.     

Passive immunization of tilapia, Oreochromis niloticus (L.), with anti-Streptococcus iniae whole sera

Passive immunization of tilapia, Oreochromis niloticus, was conducted to determine whether anti- Streptococcus iniae whole sera (ASI), heat inactivated anti- S . iniae whole sera (HIASI) and normal whole sera (NWS) were protective when intraperitoneally (i.p.) injected into tilapia. The ASI was produced in tilapia actively immunized (challenged) with virulent S. iniae by i.p. injection. An antibody response against S. iniae was demonstrated by enzyme linked immunosorbent assay (ELISA) and 18% of the immunized fish died because of the S. iniae infection. The actively immunized tilapia demonstrated a secondary antibody response and immunity to S. iniae after challenge with S. iniae by i.p. injection. Survival was 100% in the actively immunized fish. The NWS was obtained from tilapia free of ASI antibody and susceptible to S. iniae infection (40% mortality). In two separate experiments, significantly higher mortality was noted in tilapia passively immunized with NWS (33 and 53%) and phosphate buffered saline (PBS) (30 and 60%), in comparison with mortalities of 0 and 10% or 3.3 and 6.7% in the fish passively immunized with ASI or HIASI 14 days after S . iniae infection by i.p. injection ( P  = 0.0003 and 0.0023). Results suggest that immunity provided by ASI and HIASI was because of antibody against S. iniae . Inactivation of complement in the HIASI treatment further suggests that ASI antibody plays a primary role in immunity against S. iniae infection.     

Whole-body concentrations of cortisol and sex steroids in white sturgeon (Acipenser transmontanus, Richardson 1836) during early development and stress response

In general little is known about hormones and the ontogeny of the stress response in the early developmental stages of chondrostean fishes and in particular of white sturgeon (Acipenser transmontanus, Richardson 1836). In this study, we measured for the first time cortisol and sex steroids (testosterone and estradiol) in eggs, larvae, post-larvae, and fry of white sturgeon by radioimmunoassay (RIA), to elucidate some endocrine aspects of its development. The cortisol, testosterone, and 17β-estradiol of maternal origin found in unfertilized eggs of white sturgeon probably regulate both growth and development of the embryo. Cortisol decreased after fertilization, whereas testosterone and 17β-estradiol did not significantly change. During the late stages of embryo development and immediately after hatching, endogenous production of cortisol and sexual steroids, respectively, occurred. Sex steroids may be physiological inducers of gonad sex differentiation in sturgeon. All steroids showed an increase 10 days post-hatch (dph), near the transition from an endogenous to an exogenous energy source. Cortisol maintained the same basal levels even after metamorphosis, whereas testosterone and 17β-estradiol declined significantly in post-larvae at 35 and 45 days post-hatch. In addition, to evaluate the ontogeny of a functional hypothalamic-pituitary-interrenal (HPI) axis, larvae and fry were submitted to acute stress. The HPI axis did not seem to be functional on the first day post-hatch, but became so from the third day post-hatch onward.     

Case report: concurrent herpesviral and presumptive iridoviral infection associated with disease in cultured shortnose sturgeon,Acipenser brevirostrum (L.), from the Atlantic coast of Canada

Approximately 8 weeks after a chlorine insult associated with the city water supply, shortnose sturgeon, Acipenser brevirostrum (L.), from one group presented with small (3–4 mm) irregular foci of cutaneous pallor that involved the dorsocranial integument with progressive ulceration of the nascent lesions. Various bacterial organisms were isolated from the cutaneous lesions, but not from the internal viscera. Histologically, the nuclei of the intralesional and perilesional epidermal cells often exhibited margination of the chromatin that resulted in a homogenous, pale, amphophilic, tinctorial quality of the nucleoplasm consistent with a herpesvirus infection. In addition, rare lamellar epithelial cells were prominently enlarged due to an abundant, dense, basophilic cytoplasm characteristic of an iridovirus infection. Inoculation of cutaneous lesion and kidney, spleen, liver sample pools from affected shortnose sturgeon onto white sturgeon spleen (WSS‐2) cell line induced cytopathic effect characterized by syncytia formation. Ultrastructural analysis of infected WSS‐2 cells revealed viral particles with a characteristic herpesvirus morphology. Intranuclear hexagonal capsids had a diameter of 95–108 nm, and enveloped particles present in the cytoplasm of infected cells had a diameter of 176–196 nm. This is the first report of a herpesvirus and a possible iridovirus‐like infection in shortnose sturgeon.     

Effect of Rosmarinus officinalis Essential Oil and Nisin on Streptococcus iniae and Lactococcus garvieae in a Food Model System

The effects of concentrations of Rosmarinus officinalis essential oil (EO) and nisin (N) as well as temperature and storage time on growth of Streptococcus iniae and Lactococcus garvieae in Oncorhynchus mykiss were evaluated. According to analysis of the rosemary EO, the 1, 8-cineol and α-pinene were the predominant components. The growth of S. iniae was significantly decrease by EO concentrations at 4 ºC. For L. garvieae, the viable count was significantly inhibited by EO and N singly and in combinations, incubated at bath storage of 0.25 and 0.5 µg/mL proved insufficient to act against S. iniae and L. garvieae. The combinations of the rosemary EO at 0.0015% with N at 0.5 µg/mL showed stronger antimicrobial effect against two bacterial than the rosemary EO at 0% but lower than the combination with N at 0.5 µg/mL and EO at 0.045% which in turn was lower than of the rosemary EO at 0.135%. In its turn, rosemary EO showed lower antimicrobial activity than its combinations with N, which showed a bactericidal effect against the pathogens. The best inhibitory effects of EO in combinations with N for two bacterial were obtained at combinations of EO=0.135% and N=0.5 µg/mL.