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Abstract

Nontuberculous mycobacteria (NTM) have recently emerged as important bacterial pathogens of both animals and humans. In this study, we aimed to evaluate the effect of a combination of ten antibiotics with an inhibitor of efflux pumps (EPI), i.e. berberine (BER), against 6 strains of NTM. Our results showed that the BER potentiated the anti-mycobacterial activities of the antibiotics. Overall, our findings show the importance of BER in increasing the efficacy of antibiotics in NTM.
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Bibliography


Gaba S, Saini A, Singh G, Monga V (2021) An insight into the medicinal attributes of berberine derivatives: a review. Bioorg Med Chem 38: 116143.
Gentry EJ, Jampani HB, Keshavarz-Shokri A, Morton MD, Velde DV, Telikepalli H, Mitscher LA, Shawar R, Humble D, Baker W (1998) Antitubercular natural products: berberine from the roots of commercial hydrastis canadensis powder. Isolation of inactive 8-oxotetrahydrothalifendine, canadine, beta-hydrastine, and two new quinic acid esters, hycandinic acid esters-1 and -2. J Nat Prod 61: 1187-1193.
Guz L, Puk K (2022) Antibiotic susceptibility of mycobacteria isolated from ornamental fish. J Vet Res 66: 69-76.
Menichini M, Lari N, Rindi L (2020) Effect of efflux pump inhibitors on the susceptibility of Mycobacterium avium complex to clarithromycin. J Antibiot 73: 128-132.
Puk K, Guz L (2020) Occurrence of Mycobacterium spp. in ornamental fish. Ann Agric Environ Med 27: 535-539.
Song L, Wu X (2016) Development of efflux pump inhibitors in antituberculosis therapy. Int J Antimicrob Agents 47: 421-429.
Van Ingen J, Boeree MJ, Van Soolingen D, Mouton JW (2012) Resistance mechanisms and drug susceptibility testing of nontuberculous mycobacteria. Drug Resist Updat 15: 149-161.
Wang Y, Fu H, Li Y, Jiang J, Song D (2012) Synthesis and biological evaluation of 8-substituted berberine derivatives as novel an-ti-mycobacterial agents. Acta Pharm Sin B 2: 581-587.
Zhou XY, Ye XG, He LT, Zhang SR, Wang RL, Zhou J, He ZS (2016) In vitro characterization and inhibition of the interaction between ciprofloxacin and berberine against multidrug-resistant Klebsiella pneumoniae. J Antibiot 69: 741-746.
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Authors and Affiliations

K. Puk
1
L. Guz
1

  1. Department of Biology and Fish Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033 Lublin, Poland
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Abstract

Ichthyophthiriasis, which is caused by Ichthyophthirius multifiliis (Ich) infections, has a severe impact on productivity in freshwater aquaculture. These infections were previously treated effectively with malachite green, a compound that is now banned on fish farms due to its carcinogenicity. To find efficacious drugs to control Ich, flowers of tansy Tanacetum vulgare were evaluated for their antiprotozoal activity. Tanacetum vulgare extract significantly reduced the survival of Ich trophonts and theronts. In vitro, the extract killed all trophonts at 3200 mg l-1, terminated tomont reproduction at 50 mg l-1, and caused mortality of all theronts at 100 mg l-1. T. vulgare extract may be a new and efficacious drug for the control of Ich.
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Bibliography

1. Buchmann K, Sigh J, Nielsen CV, Dalgaard M (2001) Host responses against the fish parasitizing ciliate Ichthyophthirius multifiliis. Vet Parasitol 100: 105-116.
2. Fu YW, Zhang QZ, Xu DH, Xia H, Cai XX, Wang B, Liang J (2014) Parasiticidal effects of Morus alba root bark extracts against Ichthyophthirius multifiliis infecting grass carp. Dis Aquat Organ 108: 129-136.
3. Kumar V, Tyagi D (2013) Chemical composition and biological activities of essential oils of genus Tanacetum − a review. J Pharm Phytochem 2: 155-159.
4. Lahnsteiner F, Weismann T (2007) Treatment of ichthyophthiriasis in rainbow trout and common carp with common and alternative therapeutics. J Aquat Anim Health 19: 186-194.
5. Puk K, Guz L (2014) Effects of medical plant extracts on the growth of the fish parasite Spironucleus vortens. Med Weter 70: 165-168.
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Authors and Affiliations

K. Puk
1
L. Guz
1

  1. Department of Fish Diseases and Biology, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033 Lublin, Poland
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Abstract

Essential oils from plants used in traditional medicine are known as a rich source of chemically diverse compounds with specific biological activities. Achillea millefolium essential oil (AEO) was screened for in vitro activity against Babesia canis. The AEO was obtained by hydrodistillation and analysed by gas chromatography coupled to mass spectrometry (GC-MS). GC-MS revealed the presence of 47 compounds in the essential oil. Those present in the highest concentrations were chamazulene (34.45%), β-caryophyllene (8.93%), (E)-germacrene D (7.55%), patchoulene (7.27%), β-guaiene (4.62%), α-humulene (4.59%), santolina epoxide (4.41%), ethyl iso-allocholate (2.97%), aromadendrene (2.62%), and neoclovenoxid-alkohol (2.46%). AEO was found to be active in vitro against B. canis, with 50% inhibitory concentration (IC50) values of 0.06 mg/mL, as compared to imidocarb, with IC50 = 0.007 mg/mL. The study confirms that essential oil from A. millefolium has anti-babesial properties in vitro.
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Authors and Affiliations

L. Guz
1
J. Wawrzykowski
2
Ł. Adaszek
3

  1. Department of Fish Diseases and Biology, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033 Lublin, Poland
  2. Department of Animal Biochemistry, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033 Lublin, Poland
  3. Department of Epizootiology and Clinic of Infectious Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Głęboka 30, 20-612 Lublin, Poland
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Abstract

Nontuberculous mycobacteria (NTM) have recently emerged as important bacterial pathogens of animals and humans. Of particular concern is the high level of antimicrobial resistance displayed by these organisms, which complicates treatment and potential successful outcomes. Here, we evaluated the potential of Carlina acaulis L. as a source of novel anti-mycobacterial agents. Our goal was to measure the activity of aqueous, ethanol, and chloroform C. acaulis root extracts against 99 NTM strains. GC-MS spectroscopy analyses were performed to deliver qualitative and quantitative data on the composition of C. acaulis extract. In our study, we have shown for the first time the activity of C. acaulis extracts against NTM. The highest activity was exhibited by the chloroform extract, which inhibited the growth of more than 90% of the strains at the dose of 100 μg/mL (MIC90 = 100 μg/mL). The results of the GC-MS analysis of the C. acaulis chloroform extract contributed to the identification of 37 compounds, with carlina oxide as the most representative compound (69.52%) followed by 3,4-dihydro-2H-phenanthren- -1-one (6.54%) and stigmast-5-en-3-ol (4.14%). Our results indicate that C. acaulis chloroform and ethanol extracts have potential for treatment of NTM infections and that this plant contains anti-mycobacterial compounds.
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Authors and Affiliations

K. Puk
1
J. Wawrzykowski
2
L. Guz
1

  1. Department of Biology and Fish Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033 Lublin, Poland
  2. Department of Biochemistry, Faculty of Veterinary Medicine, University of Life Science in Lublin, Akademicka 12, 20-033 Lublin, Poland
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Abstract

Ichthyophthiriasis, commonly known as white spot disease, occurs in both wild and cultured fish and is responsible for heavy economic losses to the aquaculture industry. In past decade, several chemical therapeutants were used to treat ichthyophthiriasis, but the effective drugs, such as malachite green, have been banned for use in food fish due to its genotoxic and carcinogenic properties. To find efficacious drugs to control Ichthyophthirius multifiliis (Ich), whole Eclipta alba plants and dried root of Arctium lappa were evaluated for their antiprotozoal activity. E. alba and A. lappa extracts significantly reduced the survival of Ich trophonts and theronts. In vitro, the E. alba and A. lappa methanol extracts killed all trophonts at 3200 mg l-1. All trophonts were killed after exposure to E. alba aqueous extract at 3200 mg l-1. The methanol extracts of E. alba and A. lappa killed 100% of I. multifiliis theronts at 400 mg l-1 and 800 mg l-1, respectively. The aqueous extract of E. alba and A. lappa killed 100% of I. mulitifiliis theronts at 1600 mg l-1 and 3200 mg l-1, respectively. E. alba and A. lappa extracts may be new and efficacious drugs for the control of ichthyophthiriasis.
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Bibliography


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Fu YW, Zhang QZ, Xu DH, Xia H, Cai XX, Wang B, Liang J (2014) Parasiticidal effects of Morus alba root bark extracts against Ichthy-ophthirius multifiliis infecting grass carp. Dis Aquat Organ 108: 129-136.

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Puk K, Guz L (2021) Parasiticidal effects of Tanacetum vulgare extract against Ichthyophthirius multifiliis. Pol J Vet Sci 24: 159-161.

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Authors and Affiliations

P. Leśniak
1
K. Puk
1
L. Guz
1

  1. Department of Biology and Fish Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-033, Lublin, Poland
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Abstract

The aim of this study was to investigate the antimycobacterial activity of 39 free terpenes and their activity in combination with streptomycin. Antimicrobial activity was first evaluated by screening 39 free terpenes at concentrations from 1.56 to 400 μg/mL. None of these exhibited positive effects against any of the nontuberculous mycobacteria (NTM) strains tested. However, six of the 39 terpenes (isoeugenol, nerol, (+)-α-terpineol, (1R)-(−)-myrtenol, (+)-terpinen-4-ol, and eugenol) were shown to enhance the activity of streptomycin against the NTM strains isolated from diseased ornamental fish.
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Bibliography

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2. Cantrell CL, Franzblau SG, Fischer NH (2001) Antimycobacterial plant terpenoids. Planta Med 67: 685–694.
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Authors and Affiliations

L. Guz
1
K. Puk
1
D. Szwajgier
2
A. Pastuszka
1

  1. Department of Biology and Fish Diseases, Faculty of Veterinary Medicine, University of Life Sciences in Lublin, Akademicka 12, 20-033 Lublin, Poland
  2. Department of Biotechnology, Microbiology and Human Nutrition, University of Life Sciences in Lublin, Skromna 8, 20-704 Lublin, Poland
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Abstract

The in vitro anti-Babesia canis activities of nine essential oils were investigated. Among the tested essential oils Achillea millefolium, Eugenia caryophyllus and Citrus grandis were the most active (IC50 values of 51.0, 60.3 and 61.3 μg/mL, respectively). The oils from Abies sibirica, Rosmarinus officinalis, Eucalyptus globulus, Cinnamonum zeylanicum, Mentha piperita and Pinus sylvestris were less active (IC50 values of 134.3, 237.3, 239.3, 367.9, 837.5 and 907.3 μg/mL, respectively). The results support the concept that some essential oil constituents may be useful in the clinical management of babesiosis.

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Authors and Affiliations

L. Guz
J. Ziętek
K. Puk
Ł. Adaszek
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Abstract

Despite many phytochemical and pharmacological investigations, to date, there are no reports concerning the antibabesial activity of extracts of A. millefolium against B. canis. This study was aimed at investigating the biological activities of A. millefolium against the Babesia canis parasite and to identify its chemical ingredients. The water (WE), ethanol (EE) and hexane/acetone (H/AE) extracts of plant aerial parts were screened for total phenolic content (TPC), total flavo- noid compound (TFC), DPPH free radical-scavenging activity and its antibabesial activity assay. In this study, imidocarb diproprionate was used as a positive control. The H/AE and EE extracts were analysed using gas chromatography–mass spectroscopy (GC–MS).

In the EE extract, the main compounds were 17.64% methyl octadec-9-ynoate, 16.68% stigmast-5-en-3-ol(3α,24S) and 15.17% hexadecanoic acid. In the H/AE extract, the main com- pounds were 34.55% 11-decyldocosane, 14.31% N-tetratetracontane, 8.22% β-caryophyllene, and 7.69% N-nonacosane. Extract of EE contained the highest content of phenolics followed by H/AE and WE. The concentration of flavonoids in EE, H/AE and WE extracts showed that TFC was higher in the EE samples followed by H/AE and WE. The antioxidant activities were highest for AA, followed by EE, WE and H/AE. The antibabesial assay showed that the WE, EE and H/AE extracts of A. millefolium were antagonistic to B. canis. At a 2 mg/mL concentration, it showed 58.7% (± 4.7%), 62.3% (± 5.5%) and 49.3% (± 5.1%) inhibitory rate in an antibabesial assay, respectively.

Considering these results, the present findings suggest that A. millefolium extracts may be a potential therapeutic agent and that additional studies including in vivo experiments are essential.

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Authors and Affiliations

L. Guz
Ł. Adaszek
J. Wawrzykowski
J. Ziętek
S. Winiarczyk
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Abstract

Vibrio species are common inhabitants of aquatic environments and have been described in connection with fish and human diseases.
Six Vibrio species were isolated from diseased freshwater and ornamental fish in Poland. The strains were identified based on morphological and biochemical characteristics and confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) as V. albensis (n=3) from Gymnocephalus cernua, Sander lucioperca, Paracheirodon innesi, and Xiphophorus hellerii; V. mimicus (n=1) from Xiphophorus maculatus; and V. vulnificus (n=1) from Nematobrycon palmeri. This is the first time that Vibrio species have been isolated and described from ornamental fish in Poland. The isolates were resistant to ampicillin (83.3%), gentamicin (16.6%), ciprofloxacin (16.6%), sulfamethoxazole-trimethoprim (16.6%), and chloramphenicol (16.6%). The multiple antibiotic resistance (MAR) index was 0.00-0.08 for V. albensis, 0.17 for V. mimicus, and 0.33 for V. vulnificus.
Our study confirmed the presence of potentially pathogenic Vibrio species in freshwater and ornamental fish. Therefore, further monitoring of the presence of Vibrio species, mainly in ornamental fish, is necessary.
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Authors and Affiliations

A. Pastuszka
1
L. Guz
1
K. Michalak
2
D. Pietras-Ożga
2
K. Puk
1

  1. Department of Biology and Fish Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Akademicka 12, 20-950 Lublin, Poland
  2. Department of Epizootiology and Clinic of Infectious Diseases, Faculty of Veterinary Medicine, University of Life Sciences, Głęboka 30, 20-612 Lublin, Poland
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Abstract

Edible snails are kept in farms in many countries worldwide. As farm animals, they are an object of interest of veterinary studies and applied biology. There is a large demand for tests which would help identify their health and well-being. The objective of this study was to assess the usefulness of determining the concentration of urea in hemolymph as a marker of health of the Lissachatina fulica and Cornu aspersum edible snails. The observation covered snails from four farms marked from A to D, in which numerous deaths (farm A) and decreased body weight gain (farms B and C) were observed. In experimental farm D we observed a group of snails subjected to stress and a control group maintained in correct conditions. High concentrations of urea were found in the hemolymph of all farm animals from farms A, B and C, as well as in those subjected to food deprivation in farm D (on average from 96 mg/dl in farm D to 320 mg/dl in farm A). On the other hand, in controls from group D, the concentration of the parameter in question was much lower (< 2.0 mg/dl). The results obtained indicate that the urea concentration is a non-specific marker of pathological conditions in snails, and that the continuous monitoring of this parameter makes it possible to demonstrate irregularities in farming and introduce appro- priate and early measures to eliminate such disturbances.

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Authors and Affiliations

J. Ziętek
L. Guz
A. Wójcik
ORCID: ORCID
S. Winiarczyk
Ł. Adaszek
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Abstract

The aim of the study was to establish normal ranges for chosen biochemical parameters of haemolymph of snails (Gasropoda: Mollusca), in the light of the use of these animals as experi- mental models in various types of studies. The study was conducted on 100 specimens of Cornu aspersum maxima (CAM) and 100 specimens of Cepaea nemoralis (CN). The haemolymph col- lected from the animals was analysed using colorimetry to assay aspartate transaminase (AST) activity, alanine transaminase (ALT) activity, amylase activity and the concentrations of urea and triglycerides. In the further part of the study, the influence of administering doxycycline with feed on the change of AST and ALT activity in snail haemolymph has been studied. The normal values established for CAM are as follow: AST activity: 26-38 u/l, ALT activity: 0-11 u/l, amylase activity 9-16 u/l, concentration of urea: 3-6 mg/dl, concentration of triglycerides: 16-20 mg/dl. For CN, the following data have been obtained: AST activity: 30-80 u/l, ALT activity: 0-15 u/l, amylase activity 12-15 u/l, concentration of urea: 5-8 mg/dl, concentration of triglycerides: 18-24 mg/dl. It has been shown that doxycycline presents a high workload on the hepatopancreas of snails, which is reflected by a statistically significant (p<0.05) increase of AST and ALT activity in the haemolymph of the specimens which obtained doxycycline in feed, as compared to the groups with antibiotic-free feed. The haemolymph activity of both studied parameters increased together with study time and tetracycline administration time.

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Authors and Affiliations

J. Ziętek
L. Guz
S. Winiarczyk
K. Szkucik
M. Ziomek
M. Wysokowski
J. Madany
Ł. Adaszek

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