Corrigendum: Effects of dietary nutmeg (Myristica fragrans) seed meals on growth, non-specific immune indices, antioxidant status, gene expression analysis, and cold stress tolerance in zebrafish (Danio rerio).

[This corrects the article DOI: 10.3389/fnut.2022.1038748.].

Electrospinning of Nanofibers Incorporated with Essential Oils: Applications in Food.

Nowadays, modern food preservation techniques have emerged in the last decade. Recently, a combination of nanotechnology and active packaging has allowed the incorporation of bioactive compounds, such as essential oils, into nanoscale electrospun fibers. This phenomenon provides a new horizon in food safety and preservation. The incorporation of essential oils into electrospun nanofibers can extend the duration of antimicrobial and antioxidant activity of essential oils, which subsequently leads to longer shelf life, better preservation, and superior quality of food. In the current paper, the essential oils incorporated into nanofibers have been reviewed. The fabrication of nanofibers is usually carried out using different substances by applying various manufacturing methods, including needleless and needle-based electrospinning techniques. In this study, an emphasis on the antioxidant and antibacterial effects of electrospun nanofibers loaded with essential oils and their application in food models has been laid. Nevertheless, other challenges associated with using nanofibers incorporated with essential oils, such as their impact on organoleptic properties, cytotoxicity, and durability, have been discussed to achieve a holistic view of applying the electrospinning techniques in the food industry.

Application of multi-criteria decision-making for optimizing the formulation of functional cookies containing different types of resistant starches: A physicochemical, organoleptic, in-vitro and in-vivo study.

This research aimed to develop a healthy cookie formulation containing different types of resistant starch, through the application of TOPSIS approach, as a potent feature of MCDM methodologies. Physicochemical investigations reveled that a harder, denser and less sticky dough was produced by the addition of both types of RS. The baking of these doughs resulted in the production of crumblier cookies of less spread ratio, lower porous crumb and whiter surface/crumb. Moreover, in-vitro digestibility of the cookies demonstrated that the baking process can adversely reduce the resistance of RS4 to the enzymolysis reactions. This phenomenon was further corroborated by in-vivo studies where the RS4 enriched cookies were less capable in reducing the postprandial blood glucose. TOPSIS, through successful solving of the multiple criteria decision 9 (alternatives) × 15 (evaluated attributes) matrix suggested that the cookie containing 15% RS is the best alternative in all aspects, possessing acceptable physicochemical/organoleptic attributes, and in-vivo/in-vitro dietary fiber.

WITHDRAWN: Electrospinning of Nanofibers Incorporated with Essential Oils: Applications in Food

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Effects of dietary nutmeg (Myristica fragrans) seed meals on growth, non-specific immune indices, antioxidant status, gene expression analysis, and cold stress tolerance in zebrafish (Danio rerio).

Introduction: A medicinal plant, Myristica fragrans seed meal (nutmeg), was utilized to evaluate its impact on the growth, immunity, and antioxidant defense of zebrafish (Danio rerio). Methods: In this regard, zebrafish (0.47 ± 0.04 g) (mean ± S.D.) were fed with 0% (control), 1% (T1-nutmeg), 2% (T2-nutmeg), and 3% (T3-nutmeg) of powdered nutmeg for 70 days. At the end of the feeding trial, growth performance, survival rate of fish, and temperature-challenge effects were recorded. Immune and antioxidant parameters were also assessed through the collection of serum and skin mucus samples. Results: The results indicated that nutmeg supplementation did not significantly influence the growth of zebrafish (P > 0.05); however, the survival rate of fish fed with 2 and 3% of nutmeg supplementation significantly decreased (P < 0.05). The skin mucus and serum total protein, total immunoglobulin (Ig), and lysozyme activity were significantly increased in T3-nutmeg treatment in comparison to the control (P < 0.05). Superoxide dismutase (SOD) and catalase (CAT) activities were also enhanced in the T3-nutmeg group (P < 0.05). Nutmeg supplementation significantly upregulated the mRNA expression of growth hormone (gh) and insulin growth factor-1 (igf-1). Moreover, the nutmeg inclusion upregulated the expression of interleukin-1ß (IL-1ß), lysozyme, sod, and cat. The dietary supplementation of nutmeg significantly increased the resistance of zebrafish against cold-water shock and survivability afterward (P < 0.05). Discussion: In conclusion, the supplementation of 3% powdered nutmeg in zebrafish diets could be suggested as an effective immune stimulator that improves antioxidant defense and stress tolerance.

Size-dependent effects of microplastic on uptake, immune system, related gene expression and histopathology of goldfish (Carassius auratus).

Todays, with the industrialization of human societies, pollution of aquatic ecosystems with plastics derivatives are a serious concern, affecting the life of their organisms. The present study was conducted to investigate the size effects of micro-plastic, polystyrene on some physiological lesions of the goldfish, Carassius auratus. Fish were exposed to two sizes (0.25 and 8 µm) polystyrene at different environmentally relevant concentrations. The exposure trial was done in two steps. First, fish exposed to a stable concentration of 300 mg/L polystyrene for 168 h. Gill, intestine, and liver tissues were sampled every 24 h to investigate the accumulation of polystyrene. Then, fish were exposed in three replicates to 0 (control), 0.05, 0.5, and 5 mg/L polystyrene in two sizes of 0.25 and 8 µm for 28 days. After the exposure period, gill, liver, and intestine tissues were sampled for histological study, also, serum samples were collected for biochemical assays. Fluorescent microscope observations confirmed the accumulation of polystyrene in tissue samples with time. In addition, histological lesions were found in the liver, intestine, and gill of the exposed fish. The severity of lesions showed a size and dose-dependent pattern. Polystyrene induced the antioxidant system of exposed fish through elevating the levels of SOD and CAT activity and significant difference in expression of antioxidant related genes (CAT, SOD and HSP70). In conclusion, the results of the present study confirmed the toxic effects of microplastic, polystyrene on goldfish.

Effects of dietary fern (Adiantum capillus-veneris) leaves powder on serum and mucus antioxidant defence, immunological responses, antimicrobial activity and growth performance of common carp (Cyprinus carpio) juveniles.

A 56-day research was performed to examine the influence of graded levels (0 (control), 0.5, 1 and 2%) of Fern (Adiantum capillus-veneris) leaves powder (FLP) in diet on immune competence and growth of common carp (Cyprinus carpio, initial weight = 20 g). The serum total immunoglobulins content and lysozyme activity in the 1 and 2% FLP groups remarkably increased compared to the other groups (P < 0.05). The skin mucosal lysozyme activity enhanced with increasing dietary FLP level in a dose-response manner. Fish fed on the FLP-supplemented diets had higher skin-mucosal superoxide dismutase activity than the control (P < 0.05). However, serum antioxidant enzymes were not affected by dietary fern (P > 0.05). The serum bactericidal activity against human and fish pathogens increased with enhancing the FLP level in diet against Staphylococcus aureus, Escherichia coli (EHEC ATCC 43895), Escherichia coli (CI), Pseudomonas aeruginosa, Klebsiella pneumonia and Aeromonas hydrophila. The serum antibacterial activity against Yersinia ruckeri in the 2% FLP group was higher than the other treatments. Furthermore, the serum bactericidal activity against P. aeruginosa (ATCC 27853) only observed in fish fed on the 1 and 2% FLP-supplemented diets. The skin mucosal bactericidal activity and inhibitory effects increased with enhancing the FLP level in diet against E. coli, K. pneumonia, Y. ruckeri and A. hydrophila in a dose response manner. Moreover, the skin mucosal bactericidal activity against S. aureus only observed in fish fed on 1 and 2% FLP-supplemented diets. The weight gain values in the 1 and 2% FLP groups were higher than the other treatments (P < 0.05). Feed conversion ratio (FCR) improved with increasing FLP level in diet in a dose-response manner (P < 0.05). By considering serum and mucosal bactericidal activities against different pathogenic bacteria, the supplementation of 2% FLP in diet is recommended for C. carpio during grow-out phase.

Dietary supplementation of lemon verbena (Aloysia citrodora) improved immunity, immune-related genes expression and antioxidant enzymes in rainbow trout (Oncorrhyncus mykiss).

The use of veterinary drugs for the treatments of fish diseases has often health and environmental side effects and thus alternative more sustainable strategies are needed. In this study we have explored the effect of dietary administration of lemon verbena (Aloysia citrodora) leaves powder on growth, immune parameters and antioxidant enzyme activity of rainbow trout (Oncorrhyncus myskiss). The results show that adding lemon verbena (LV) leaves powder (0.5, 1 and 2%), as feed supplement does not affect significantly rainbow trout growth, but increases the levels of some immune parameters. All enriched-diets induced an increase in the levels of lysozyme activity and total immunoglobulin in rainbow trout serum (P < 0.05), whilst 1 and 2% enriched-diets increased lysozyme and immunoglobulin in skin mucus. Expression of interleukin-1ß (IL-1ß) was enhanced in all fish supplemented with LV leaves powder (0.5, 1 and 2%) whilst only the diet with the highest dose (2%) induced a significantly higher expression of IL-8 and tumour necrosis factor (TNF-α). Fish fed with LV leaves powder at 1 and 2% had significantly lower expression of transforming growth factor (TGF-ß, P < 0.05). Furthermore, all enriched-diets significantly increased the activity of super oxide dismutase (SOD, P < 0.05) and diets containing 1 and 2% of LV leaves powder also increased activity of glutathione-S-transferase (GST) and glutathione peroxidase (GPx). In conclusion, LV leaves powder administered as feed supplement at 2% enhances the expression of some immune-related genes (IL-1ß, IL-8 and TNF- α), rises the levels of lysozyme and total immunoglobulin in serum and skin mucus and increases the activity of antioxidant enzymes (SOD, GST and GPx) in rainbow trout.

Rainbow trout fillet biopreservation by edible chitosan-based coating containing egg yolk antibody (IgY) and lycopene.

The aim of the study was to investigate the effect of the extracted egg yolk antibody along with lycopene on the chemical quality of the rainbow trout fillet during 16 days of refrigeration storage. Chickens were immunized against Pseudomonas fluorescens (P. fluorescens), Shewanella putrefaciens (S. putrefaciens) and total spoilage bacteria and their eggs were collected for the isolation of egg yolk antibodies. Then fish fillets were immersed in chitosan-based coating solutions, containing lycopene and extracted antibodies, and analyzed for lipid oxidation changes (peroxide, thiobarbituric acid, free fatty acid and fatty acid profile), physico-chemical properties (pH and water holding capacity), and sensory evaluation, during 16 days of refrigeration storage. Results showed that chitosan solutions with lycopene or IgY could significantly (p < 0.05) increase the oxidative stability of lipids in fish fillets; although, combinational use of lycopene and IgY showed a higher effect on delaying the rate of lipid oxidation. Significant differences were also observed between treatments contained combination of chitosan, antibody and lycopene with the control group, regarding pH and WHC. Saturated fatty acids increased in all treatments, although the changes in the treatments containing lycopene and antibody were significantly (p < 0.05) lower than the control group. Hence the addition of egg yolk antibody and lycopene in coating solution are good bio-preservatives for seafood products as it improves sensory attributes and prevents lipid oxidation.

Effect of walnut kernel septum membranes hydroalcoholic extract on the shelf life of traditional butter.

Incorporation of natural ingredients antioxidants in edible fats can profitably affect their oxidative stability during production and storage. The purposes of the current work were to assess the antioxidant and antimicrobial effect of walnut kernel septum membranes hydroalcohol extract (WHE) in traditional butter (TB). Antioxidant characterization of the extract was screened through methods of DPPH, reducing power and total phenolic assays. After preparation of traditional butter from yogurt, WHE was incorporated into TB at three different concentrations; 0.05%, 0.1% and 0.5% and compared with a control, BHT and tocopherol treated samples (200 mg of BHT and tocopherol/kg). Microbiological studies (Staphylococcus aureus, Coliforms, Psychrotrophic bacteria, yeasts and molds) were done during 90 days of storage time. Changes in Anisidine value (AV), acid value, peroxide value (PV) free fatty acids (FFA), Schaal and Totox value were monitored at 45-day intervals. Sensory evaluation was done using 10 semi-trained panelists based on the 5-point hedonic scale. It was found that the total phenolic content of WHE was 368.86 mg GAE/g. The BHT had higher antioxidant activity than WHE inhibiting 92.3% of the DPPH radical at 600 µg/mL. Peroxide value of TB treated with tocopherol, BHT and WHE 0.5% was 0.29 ± 0.07, 0.39 ± 0.07 and 0.52 ± 0.04 respectively. Furthermore, the WHE incorporated butter has shown low levels of free fatty acids, Schaal and Totox value when compared to control treatment. The WHE 0.5% incorporated sample had the most antimicrobial activity and it inhibited the growth of all the microorganisms (except Staphylococcus aureus) used in the study. Among the treated TB, the samples treated with the control and WHE 0.05% had the highest sensory attributes score. The study showed that WHE could be an excellent natural origin of antimicrobial and antioxidant agents which can be used in butter.

Physicochemical and morphological properties of resistant starch type 4 prepared under ultrasound and conventional conditions and their in-vitro and in-vivo digestibilities.

In the present work, cross-linked resistant starch (RS4) was prepared under sonication and conventional conditions at various levels of pH (9-12) and cross-linker concentration (sodium trimetaphosphate/sodium tripolyphosphate 99:1, 5-15%). It was found that phosphorous and resistant starch content was generally increased by increasing the cross-linker concentration, pH and application of sonication. The damage to the surface of sonicated granules was revealed by scanning electron micrographs. The presence of cross-linked phosphorous groups was demonstrated by FT-IR results through the appearance of a new peak at wave numbers of 1248-1252 cm-1 that was more conspicuous in sonicated cross-linked samples. Sonicated cross-linked starches showed higher gelatinization temperatures and lower degrees of crystallinity, while no changes was detected in terms of A-type crystalline pattern. The development of viscosity was diminished prominently by the extreme cross-linking reactions in both sonicated and conventional cross-linked starches. The least glycemic index value was obtained for sonicated cross-linked starches which was negatively correlated to their higher RS content measured in-vitro. These results provide novel information on the preparation of cross-linked resistant starch under sonication conditions.

Characterization of Gamma-Irradiated Rosmarinus officinalis L. (Rosemary).

OBJECTIVES: Rosmarinus officinalis L., a member of the family Lamiaceae, is regarded as the spice with the highest antioxidant activity. MATERIALS AND METHODS: In this study, the transmission electron microscopy, X-ray diffraction, and fourier transform infrared spectroscopy (FTIR) physicochemical characteristics of the nanostructure of gamma-irradiated rosemary were investigated. RESULTS: The particle size distribution of the gamma-irradiated rosemary prepared under irradiation at 30 kGy in a Cobalt-60 irradiator exhibited a very narrow size distribution with average size of 70 nm. The results showed that irradiated (30 kGy) and crude rosemary had similar patterns of FTIR spectra, typical of phenol compound, without any notable changes in the key bands and functional groups status. Rosemary irradiated with 50 kGy and 10 kGy showed the highest and lowest crystallinity, respectively. Rosemary crystallinity of irradiated samples was lower compared with the nonirradiated sample. CONCLUSION: Therefore, 30 kGy can be optimum for the synthesis of nanoparticles, average size of 70 nm, with low crystallinity and without any notable change in key bands compared to nonirradiated samples.

Health Risk Assessment for Human Exposure to Trace Metals and Arsenic via Consumption of Hen Egg Collected from Largest Poultry Industry in Iran.

The relative contribution of foodstuffs to intake of heavy metal is still equivocal, and thus, available data are rare. Here, the concentration of ten heavy metals and arsenic were measured in the content of hen egg from five strains of commercial laying hens to evaluate contamination level and potential health risk among local and international consumers in Iran. The estimated daily intake (EDI) of all heavy metals was significantly lower than the provisional tolerable daily intake (PTDI) and respective tolerable daily intake (TDI). The non-carcinogenic and carcinogenic risk values for eating eggs were in the safe range for all consumers, indicating that the no health risk for consumers due to the daily intake of either arsenic or heavy metals. Therefore, intake of heavy metal exposure through the consumption of commercial egg product in Iran is completely safe for both national and international consumers.

Using WPC-inulin-fucoidan complexes for encapsulation of fish protein hydrolysate and fish oil in W1/O/W2 emulsion: Characterization and nutritional quality.

The double emulsions and freeze-dried microcapsules containing fish protein hydrolysate (FPH) and fish oil (FO) were stabilized by complexs of whey protein concentrate (WPC) with inulin (Inu) and fucoidan (Fuc) in terms of physical characteristics (particle size distribution, morphology, encapsulation efficiency, solubility, …), oxidative stability, nutritional quality and in vitro release. Higher encapsulation efficiency and solubility were observed in Fuc-WPC microcapsules (86.31% and 30.26 mg/100 g, respectively). The combination of Fuc-WPC in the wall material showed higher oxidative stability than other wall material. The higher values of PUFA and SFA were observed in Inu-WPC Fuc-WPC microcapsules, respectively. The Fuc-WPC and Inu + Fuc-WPC micrographs showed a more porous structure compared to Inu-WPC. The mean particle size ranged from 536.8 ±â€¯52.70 to 842.36 ±â€¯21.41 nm. No significant differences were observed in the released oil and the fatty acid composition during gastrointestinal digestion. Sensory evaluation of fortified natural yogurt with microcapsules showed lower fishy flavor in Inu-WPC samples than those fortified with Fuc-WPC and Inu + Fuc-WPC. In general, the use of inulin with WPC as a wall materal resulted in good characteristics and sensory attributed, although the use of fucoidan with WPC conferred higher oxidative stability during storage.

The impact of strain and feed intake on egg toxic trace elements deposition in laying hens and its health risk assessment.

The impact of strain or feed intake on food trace elements and its health risk assessment is still ambiguous, and therefore, available facts are rare. Therefore, this study aimed to assess the effects of both strains and feed intake on trace elements depositions to egg, toxic heavy metals concentration, and health risk assessment of egg consumption. In the current cross-sectional study, the selected strains, including Shaver White, Hy-Line W36, Bovanse White, Lohman LSL-Lite, and Native laying hens of Khorasan Razavi province, were examined. A total number of 50 samples of eggs and 15 samples of their feed was purchased from poultry farms. Yolk and white were separately analyzed. Sample preparation was performed by wet digestion followed by Inductively Coupled Plasma Mass Spectrometry. Trace metals Pb, As, Cd, Hg, Cr, and Ni were detected. Statistical analysis was performed in Stata11.2 portable software. Although there was a significant difference in strains and feed, no significant difference was observed in trace elements in egg contents (weight of egg white and egg yolk). Mercury concentration in all the samples was below the instrument detection limit. In this study, the target hazardous quotients were below one for all trace elements. Therefore, Iranian does not experience the adverse health effects due to the consumption of egg.

Prevalence of liver flukes infections and hydatidosis in slaughtered sheep and goats in Nishapour, Khorasan Razavi, Iran.

BACKGROUND: Food-borne trematode infections and hydatidosis are endemic diseases caused by helminths in Iran that are responsible for great economic loss and getting public health at risk. AIM: Aim of this study was to determine the prevalence of fasciolosis, dicrocoeliasis, and hydatidosis infections in slaughtered sheep and goats in Nishapour, Khorasan Razavi province of Iran. MATERIALS AND METHODS: A survey was implemented on 130,107 sheep and goats slaughtered at an abattoir in Nishapour (Neyshbur) city, north central Khorasan Razavi Province, Iran, to determine the prevalence of fascioliasis, dicrocoeliosis and presence of hydatidosis. RESULTS: During a 1-year period of study, among 130,107 of sheep and goats slaughtered at Nishapour abattoir, 1064 and 7124 livers were condemned totally and partially, respectively. A total of 255 (0.19%), 181 (0.12 %), and 7751 (5.95%) of livers were condemned due to cysts of Echinococcus granulosus, flukes of Fasciola spp., and Dicrocoelium dendriticum, respectively. Totally, 1932 (1.48%) lungs were condemned due to hydatidosis. The significant seasonal pattern was seen for fasciolosis, dicrocoeliosis, and hydatidosis, statistically (p<0.01). CONCLUSION: According to this study, it seems that Neyshabour is considered as an endemic region for Fasciola spp. and D. dendriticum infections and D. dendriticum is the most widespread liver fluke found in sheep and goats.

An investigation of the effect of Zataria multiflora Boiss and Mentha piperita essential oils to improve the chemical stability of minced meat.

BACKGROUND AND AIM: Extending the shelf life of foods is an essential concept in food safety. Most of the time, foods deteriorate through the growth of microorganisms or oxidation process. Essential oils (EOs) derived from plant material have well-documented antioxidant and antibacterial activity. This study aimed to evaluate the effect of Zataria multiflora Boiss EO (ZEO) and Mentha piperita EO (MEO) on the chemical stability of minced meat during storage at 7°C. MATERIALS AND METHODS: Total phenolic content, ß-Carotene bleaching test, ferric reducing antioxidant potential assay, and 2,2-Diphenyl-1-picrylhydrazyl radical scavenging activity were used to determine the antioxidant potential of EOs. Five different groups including control, ZEO 0.3%, ZEO 0.5%, MEO 0.3%, and MEO 0.5% were designed to assess the chemical stability of minced meat by measuring pH, thiobarbituric acid (TBA), total volatile base nitrogen (TVBN), and peroxide value (PV). RESULTS: pH did not have any significant change during storage. TBA values in the control group were significantly higher than the treatment groups, especially from the 5th day of storage. TVBN in the treatment group was significantly lower than the control group during storage. PV values in the treatment group were significantly lower than the control group during storage. CONCLUSION: Results indicate that ZEO and MEO had an excellent antioxidant activity and retarded the spoilage process in minced meat.

Modifying attitude and intention toward regular physical activity using protection motivation theory: a randomized controlled trial.

We were guided by the Protection Motivation Theory to test the motivational interviewing effects on attitude and intention of obese and overweight women to do regular physical activity. In a randomized controlled trial, we selected using convenience sampling 60 overweight and obese women attending health centres. The women were allocated to 2 groups of 30 receiving a standard weight-control programme or motivational interviewing. All constructs of the theory (perceived susceptibility, severity, self-efficacy and response efficacy) and all anthropometric characteristics (except body mass index) were significantly different between the groups at 3 study times. The strongest predictors of intention to do regular physical exercise were perceived response efficacy and attitude at 2- and 6-months follow-up. We showed that targeting motivational interviewing with an emphasis on Protection Motivation Theory constructs appeared to be beneficial for designing and developing appropriate intervention to improve physical activity status among women with overweight and obesity.

Prevalence and antibiotic resistance pattern of bacteria isolated from urinary tract infections in Northern Iran.

BACKGROUND: This study aimed to investigate the bacteria associated with urinary tract infection (UTI) and antibiotic susceptibility pattern of the isolates during 2013-2015 in Northern Iran. MATERIALS AND METHODS: Overall 3798 patients with clinical symptoms of UTI were subjected as samples, and they were cultured and pure isolated bacteria were identified using biochemical tests and subjected to antibiogram assessment using disc diffusion method. RESULTS: Totally, 568 (14.96%) from 3798 patients had positive UTI. Four hundred and ninety-seven (87.5%) from 568 isolated bacteria were resistant to at least one antibiotic. Escherichia coli, Staphylococcus spp., and Pseudomonas spp. were the most prevalent bacteria. Isolated bacteria indicated the highest antibiotic resistance to methicillin (76.06%) and ampicillin (89.29%) and also revealed the most sensitivity to imipenem (99.1%) and amikacin (91.57%). Statistical analysis of the resistance pattern trend during 3 years indicated the insignificant increase (P > 0.05) in antibiotic resistance of the isolates. CONCLUSION: The results of this study revealed a great concern for emerging UTI-related multidrug-resistant strains of bacteria causing UTI in Iran.

Antimicrobial Resistance and Virulence-Associated Genes of Campylobacter spp. Isolated from Raw Milk, Fish, Poultry, and Red Meat.

This study was designed and conducted to evaluate the frequency, antimicrobial resistance, and presence of six virulence-associated genes among thermophilic Campylobacters isolated from raw milk, poultry (chicken, turkey, and duck), fish, cattle, and sheep meat. Out of 590 samples, which were recovered from different origins, 141 (23.9%) samples were positive for Campylobacters. Campylobacter spp. was isolated in 40.8% (106/260), 14% (28/200), and 8.7% (7/80) of poultry meat, red meat, and milk samples, respectively. Antimicrobial susceptibility test indicated a high frequency of resistance to ciprofloxacin, tetracycline, and nalidixic acid among the isolates. Furthermore, prevalence of waaC, ciaB, and pldA genes were 91.7%, 86.7%, and 80.8%, respectively; and, none of the isolates harbored both wlaN and cgtB genes, simultaneously. Moreover, there was a weak correlation between antibiotics resistance and presence of the pathogen genes. However, the existence of Campylobacter spp. isolates in food animal products, with high resistance to antibiotics and several virulence gene possessions, is alarming and increases the attention to the widespread use of antibiotics.