In vitro Genotoxicity and In silico Docking Analyses of the Essential Oils of Thuja orientalis.

Two main objectives were pursued to assess the reliability of Thuja orientalis essential oils (TOEO). The first objective was to extract TOEO, analyze them by GC-MS, and determine their in vitro genotoxicity against selected plants using the RAPD-PCR method. The second objective was to evaluate the in-silico toxicity of TOEO. The binding sites and energies of each content was calculated against B-DNA. In-silico analyses were performed using a simulation program, AutoDock Vina, and Toxicity Estimation Software Tools. 3-carene, cedrol, and 2-pinene were identified as the predominant components. In vitro studies showed that the TOEO had a more significant impact on reducing genomic stability in wheat compared to the amaranth. The lowest stability was determined as 39.78 % in wheat and 53.58 % in amaranth. Cedrol (-5,7 kcal/mol) and selinene (-5,6 kcal/mol) exhibited the highest binding affinity. The toxicity test indicated that components other than cyclohexene may have toxic effects, none of them were predicted to be mutagenic, and LD50 (mol/kg) values could vary between 1.33 and 1.55.

Limited genetic variability and spatial population structure in grasshoppers between natural and metal-contaminated areas in Egypt.

Pollutants in an environment can have long-term implications for the species living there, resulting in local adaptations with implications for their genetic structure. Heavy metal pollutants infiltrate soils and groundwater, bioaccumulate in food webs, and negatively impact biota. In this study, we investigated the degree to which the genetic structure and variability of the slender green-winged grasshopper (Aiolopus thalassinus (Fabricius) (Orthoptera: Acrididae)) were impacted by heavy metal pollution and distance. We used the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method to examine the genetic variability of populations in 3 heavy metal-polluted and 3 unpolluted locations across varying geographical distances in Egypt. The heavy metal concentrations of cadmium, copper, lead, and zinc were measured from the grasshopper tissue and soils. Sixty-nine unique and polymorphic bands were produced by 4 primers. Cluster and principal component analyses separated the populations inside and outside Cairo into 2 main branches, which were further divided into smaller branches corresponding to their geographical regions. We found no differences in the Shannon genetic diversity index between populations or with increasing heavy metal concentrations in either the soil or the grasshopper tissue. Our results showed a greater genetic variation among populations than between populations within the same location, indicating populations within locations were less differentiated than those between locations. The moderate correlation between genetic similarity and spatial distance suggests geographical isolation influenced grasshopper population differentiation. Based on the RAPD analysis, environmental pollutants and geographical distances impact the A. thalassinus population structure, potentially restricting gene flow between sites even at small spatial scales.

Morphological and molecular comparison as a useful tool for identification of the three centric marine diatoms (Bacillariophyceae: Chaetoceros).

The objective of this study was to identify morphological and molecular comparison of three marine Chaetoceros species using microscopic observations, sequence analysis of 18S rDNA, random amplified polymorphic DNA (RAPD-PCR) barcoding and nuclear magnetic resonance (NMR) spectroscopy. Chaetoceros were obtained from three different algae laboratories: Center of Excellence for Marine Biotechnology (CEMB), Chanthaburi Coastal Fisheries Research and Development (CHAN) and Institute of Marine Science, Burapha University (BIM). Genomic DNA for the RAPD-PCR analysis was extracted using the phenol-chloroform method, followed by 18S rDNA amplification. The blast results of 18S rDNA sequence confirmed the significantly matched to C. gracilis for Chaetoceros BIM and CHAN and C. muelleri for Chaetoceros CEMB(e-value = 0.0, identity = 99%). The RAPD-PCR results revealed differences in the three Chaetoceros isolates with polymorphisms between 30.43% and 60.00%, and Chaetoceros CEMB showed high polymorphic bands. Scanning electron microscopy revealed that Chaetoceros CEMB were larger and had larger setae compared to the other isolates (P < 0.05). The results of the NMR characterization of metabolites were consistent with the results of the sequence and morphological analyses. The concentrations of several metabolites, including chlorophyll c1, chlorophyll a, Myo-inositol, fucoxanthin, astaxanthin, lutein and zeaxanthin, were lower in Chaetoceros CEMB than in Chaetoceros BIM and CHAN. However, high concentrations of fatty acids, such as oleic acid, linoleic acid, α-linolenic acid and arachidic acid, were observed in all isolates. Generally, the results of this study will aid future studies examining the diversity of Chaetoceros in various cultural environments.

Legionella anisa or Legionella bozemanii? Traditional and molecular techniques as support in the environmental surveillance of a hospital water network.

Understanding the actual distribution of different Legionella species in water networks would help prevent outbreaks. Culture investigations followed by serological agglutination tests, with poly/monovalent antisera, still represent the gold standard for isolation and identification of Legionella strains. However, also MALDI-TOF and mip-gene sequencing are currently used. This study was conducted to genetically correlate strains of Legionella non pneumophila (L-np) isolated during environmental surveillance comparing different molecular techniques. Overall, 346 water samples were collected from the water system of four pavilions located in a hospital of the Apulia Region of Italy. Strains isolated from the samples were then identified by serological tests, MALDI-TOF, and mip-gene sequencing. Overall, 24.9% of water samples were positive for Legionella, among which the majority were Legionella pneumophila (Lpn) 1 (52.3%), followed by Lpn2-15 (20.9%), L-np (17.4%), Lpn1 + Lpn2-15 (7.1%), and L-np + Lpn1 (2.3%). Initially, L-np strains were identified as L. bozemanii by monovalent antiserum, while MALDI-TOF and mip-gene sequencing assigned them to L. anisa. More cold water than hot water samples were contaminated by L. anisa (p < 0.001). PFGE, RAPD, Rep-PCR, and SAU-PCR were performed to correlate L. anisa strains. Eleven out of 14 strains identified in all four pavilions showed 100% of similarity upon PFGE analysis. RAPD, Rep-PCR, and SAU-PCR showed greater discriminative power than PFGE.

Study of antibiotics sensitivity pattern and molecular characterization of Staphylococcus aureus isolated from human and animal pyogenic cases.

Staphylococcus aureus has been described as the most common cause of human and animal diseases and has emerged as a superbug due to multidrug resistance. Considering these, a total of 175 samples were collected from pyogenic cases of humans (75) and animals (100), to establish the drug resistance pattern and also for molecular characterization of human and animal isolates. Thermonuclease (nuc) gene amplification was used to confirm all presumptive S. aureus isolates and then, antibiotic sensitivity and slide Coagulase tests were used for phenotypic characterization of isolates. Following that, all the isolates were subjected to PCR amplification to detect the existence of the Methicillin-resistant (mecA) and Coagulase (coa) genes. Lastly, typing was done using the Randomly Amplified Polymorphic DNA-PCR. The overall prevalence of S. aureus in human and animal samples was found to be 39.4%. Drug sensitivity revealed the highest resistance against the ß-lactam antibiotics such as ampicillin (94.8%) and penicillin (90.6%), followed by cephalosporin (cefixime-67.7%) and quinolone (ciprofloxacin-52.1%) group of drugs. The drug sensitivity was the highest against antibiotics like chloramphenicol (95%) followed by gentamicin (90%). Among the 69 S. aureus isolates, the overall presence of MRSA was 40.5% (27.5% and 50% in human and animal isolates, respectively). Total 33 isolates exhibited coa genes amplification of more than one amplicons and variable in size of 250, 450, 800, and 1100 bp. The RAPD typing revealed amplification of five and six different band patterns in humans and animals, respectively, with two common patterns suggesting a common phylogenetic profile.

Genotypic and phenotypic diversity of Prototheca spp. recovered from bovine mastitis in terms of antimicrobial resistance and biofilm formation ability.

BACKGROUND: The Prototheca algae have recently emerged as an important cause of bovine mastitis globally. Isolates from bovine mastitis in several countries were nearly all identified as P. bovis, suggesting that it was the main causative agent of bovine protothecal mastitis. The aim of the present study was to evaluate the presence and isolation of Prototheca spp. in dairy farms, detect the genetic diversity among strains, determine the capacity of producing biofilm and their resistance to antifungal and antimicrobial drugs. RESULTS: A total of 48 Prototheca isolates from four different farms were randomly selected to be investigated. Multiplex PCR showed all isolated colonies were Prototheca bovis. Performing RAPD-PCR by using OPA-4 primer, it was revealed that there was a clear amplification pattern. Different levels of biofilm production were observed among strains. Among 48 isolates, only 4 of them (8.33%) showed strong biofilm production. By using E-test strips, amphotericin B was able to inhibit the growth of all the strains tested. Disc diffusion method used for antimicrobial sensitivity test showed that the highest activity was demonstrated by gentamicin and colistin with 95.83% (46/48) and 89.58% (43/48) of sensitive strains, respectively. CONCLUSIONS: The present study showed that RAPD-PCR was a rapid tool for discriminating P. bovis strains. Also, gentamicin and colistin can be considered as potential antimicrobial drugs which can prevent the growth of the mentioned strains in vitro, although there is no effective clinical treatment yet. Further studies are needed in order to detect an effective clinical therapy considering biofilm production by Prototheca spp. and their probable role in Prototheca pathogenicity.

Molecular identification and evaluation of gamma irradiation effect on modulating heavy metals tolerance in some of novel endophytic fungal strains.

Heavy metal (HM) pollution is a worldwide environmental issue. Given the urgent need to develop more powerful approaches for effective phytoremediation of HMs, isolation of novel endophytic strains from hyperaccumulator plants having potent HM tolerance is the main objective in this research. Moreover, the recovered strains were characterized and subjected to radiation mutagenesis to enhance their tolerance to HMs. Among 105 isolates, Alternaria alternata AUMC14431 was identified as the most effective Cd+2 tolerant strain having high recorded tolerance index (TI) (76.24%); in addition, the recorded minimum inhibitory concentration (MIC) was 300 ppm. Meanwhile, Chaetomium globosum AUMC14432 was identified as the most effective Pb+2 and Ni+2 tolerant strain having high recorded TI (97.46 and 93.34%, respectively); in addition, the evaluated MICs were 250 and 200 ppm, respectively. UV and gamma irradiation of the tested strains enhanced their Cd+2 and Pb+2 tolerance significantly (P ≤ 0.05). Meanwhile, irradiation had a negative impact on Ni+2 tolerance of C. globosum. The mutation incidence at the molecular level arising from exposure to irradiation was investigated. Genomic DNA of both the wild and mutated endophytic strains were isolated followed by random amplified polymorphic DNA (RAPD-PCR) analysis, using two short primers. A remarkable difference in DNA gel pattern between the wild type and mutated strains was observed. In conclusion, the novel isolated and irradiated endophytic strains, A. alternata S5 and C. globosum El26, having high efficiency in Cd+2 and Pb+2 tolerance, respectively, are considered to be prospective and powerful bioremediation candidates for potential application in microbially assisted phytoremediation.

Biodiversity of yeasts isolated during spontaneous fermentation of cool climate grape musts.

Biodiversity of native yeasts, especially in winemaking, has hidden potential. In order to use the value of non-Saccharomyces strains in wine production and to minimise the possibility of its deterioration, it is necessary to thoroughly study the yeast cultures present on grape fruits and in grape must, as well as their metabolic properties. The aim of the study was to characterise the yeast microbiota found during spontaneous fermentation of grape musts obtained from grape varieties 'Rondo', 'Regent' and 'Johanniter'. Grapes from two vineyards (Srebrna Góra and Zadora) located in southern Poland were used for the research. Succession of subsequent groups of yeasts was observed during the process. Metschnikowia pulcherrima yeasts were identified both at the beginning and the end of the process. Hanseniaspora uvarum, Wickerhamomyces onychis and Torulaspora delbrueckii strains were also identified during the fermentation. Torulaspora delbrueckii and Wickerhamomyces onychis strains were identified only in grape musts obtained from grapes of the Zadora vineyard. These strains may be characteristic of this vineyard and shape the identity of wines formed in it. Our research has provided specific knowledge on the biodiversity of yeast cultures on grapes and during their spontaneous fermentation. The research results presented indicate the possibility of using native strains for fermentation of grape musts, allowing to obtain a product with favourable chemical composition and sensory profile.

Antibacterial effects of Octenicept, and benzalkonium chloride on Acinetobacter baumannii strains isolated from clinical samples and determination of genetic diversity of isolates by RAPD-PCR method.

BACKGROUND: Acinetobacter baumannii (A. baumannii) is among the important causes of nosocomial infections. Due to the emergence of antibiotic resistance, many problems have been raised in the successful treatment of patients infected by this bacterium with the subsequent mortality. Therefore, the present study was performed to evaluate the antibacterial effect of Octenicept (OCT), and Benzalkonium chloride (BZK) against A. baumannii strains isolated from clinical samples, and to determine the genetic diversity of strains by RAPD-PCR method. METHODS: A total of 119 A. baumannii isolates were collected and confirmed by conventional culture and biochemical tests and PCR assay. Susceptibility of the isolates to antibiotics was evaluated by standard antibiotic susceptibility testing (AST). For antiseptics OCT and BZK, Minimum inhibitory concentration (MIC) was assessed by broth microdilution method. The prevalence of qacE and qacΔE1 genes related to antiseptics was estimated by PCR assay. Finally, genetic diversity of strains was determined by using RAPD-PCR. RESULTS: All 119 suspected isolates were confirmed as A. baumannii using conventional microbiologic tests and PCR assay. The isolates were mostly originated from blood samples. In AST, the lowest resistance was seen for ciprofloxacin and gentamicin. For antiseptics, the MIC values were reported as 15.26 µg/ml for OCT and 640 µg/ml for BZK. The antiseptic genes of qacE and qacΔE1 were found to be present in 56 (47.05%) and 59 (49.57%) of isolates respectively. RAPD typing revealed great diversity among A. baumannii isolates, with 37 clusters in isolates from ICU, of which 32 clusters were single and 5 were multiple. CONCLUSIONS: Considering the increase of resistance to antiseptics, it is of importance to monitor the susceptibility of A. baumannii to antiseptics and to promote antiseptic stewardship in hospitals. Furthermore, in this study great diversity was observed among A. baumannii isolates, which is important in understanding the molecular epidemiology of the outbreaks caused by this organism in the hospitals.

Significant contribution of the CmeABC Efflux pump in high-level resistance to ciprofloxacin and tetracycline in Campylobacter jejuni and Campylobacter coli clinical isolates.

BACKGROUND: Campylobacter resistance to antimicrobial agents is regarded as a major concern worldwide. The aim of this study was to investigate the expression of the CmeABC efflux pump and the RAPD-PCR pattern in drug-resistant Campylobacter isolates. METHODS: A total of 283 stool specimens were collected from children under the age of five with diarrhea. The minimum inhibitory concentration (MIC) of tetracycline and ciprofloxacin was determined by broth microdilution method and E-test, respectively. Detection of tetracycline and ciprofloxacin determinants was done by amplification of tetO gene and PCR-sequencing of the gyrA gene. The cmeABC transcriptional expression was analyzed by Real-time (RT)-PCR. Clonal correlation of resistant strains was determined by RAPD-PCR genotyping. RESULTS: Out of 283 fecal samples, 20 (7.02%) samples were positive for Campylobacter spp. Analysis of duplex PCR assay of the cadF gene showed that 737 and 461 bp amplicons were corresponding to Campylobacter jejuni and Campylobacter coli, respectively. All of the 17 phenotypically tetracycline-resistant Campylobacter isolates harbored the tetO gene. Also, four phenotypically ciprofloxacin-resistant Campylobacter isolates had a point mutation at codon 257 of the gyrA gene (ACA to ATA; Thr > Ile). High-level expression of the cmeA gene was observed in ciprofloxacin-resistant and high-level tetracycline-resistant Campylobacter isolates, suggesting a positive correlation between the cmeA gene expression level and tetracycline resistance level. Moreover, a statistically significant difference was observed in the cmeA gene expression between ciprofloxacin-resistant and ciprofloxacin-susceptible strains, which signifies the crucial contribution of the efflux pump in conferring multiple drug resistance phenotype among Campylobacter spp. RAPD analysis of Campylobacter isolates exhibited 16 different patterns. Simpsone`s diversity index of RAPD-PCR was calculated as 0.85, showing a high level of homogeneity among the population; however, no clear correlation was detected among tetracycline and/or ciprofloxacin resistant isolates. CONCLUSION: Significant contribution of the CmeABC efflux pump in conferring high-level resistance to tetracycline and ciprofloxacin was observed in C. jejuni and C. coli clinical isolates. The resistant phenotype is suggested to be mediated by CmeABC efflux pumps, the tetO gene, and point mutation of the gyrA gene. Genotyping revealed no clonal correlation among resistant strains, indicating distinct evolution of tetracycline and ciprofloxacin resistant genotypes among the isolates.

Prevalence of ß-lactamase-encoding genes and molecular typing of Acinetobacter baumannii isolates carrying carbapenemase OXA-24 in children.

BACKGROUND: ß-Lactam antibiotics have been broadly used for the treatment of Acinetobacter baumannii infections, resulting in development of ß-lactam inactivating ß-lactamases. Here, we described antibiotic resistance rate, prevalence of ß-lactamase-encoding genes, and clonal relationships of A. baumannii strains isolated from children referred to Children's Medical Center in Tehran, Iran, during 2019-2020. METHODS: A total of 60 non-replicate A. baumannii isolates were recovered from clinical specimens of pediatric patients. Antibiotic susceptibility testing was done by the disc diffusion method. Colistin susceptibility of isolates was performed by the broth microdilution method. ß-lactamase-encoding genes were characterized by PCR. The presence of ISAba1 element upstream of the several oxacillinase genes was also checked. Genetic relatedness of isolates was determined by using random amplification of polymorphic DNA (RAPD) typing. RESULTS: The antimicrobial susceptibility tests showed that 83.3% of A. baumannii isolates were MDR, and 40% XDR. Both MDR and XDR A. baumannii isolates were susceptible to colistin. The frequency of blaOXA-51-like, blaOXA-23-like, blaTEM, blaOXA-24-like, blaPER, blaSHV, blaCTX-M, blaOXA-58-like, and blaIMP was 100, 93.33, 60, 36.67, 28.33, 8.33, 5, 3.33, and 1.67%, respectively. Coexistence of ISAba1/blaOXA-23-like and ISAba1/blaOXA-51-like was observed in 65% and 85% of isolates, respectively. RAPD analysis revealed 4 common types and 2 single types of A. baumannii isolates. CONCLUSIONS: The multiple clones harboring blaOXA-23-like, ISAba1-blaOXA-51-like, and ISAba1-blaOXA-23-like were responsible for the spread of A. baumannii isolates in our clinical wards. Dissemination of the well-established clones is worrisome and would become therapeutic challenges due to the possible transferring genetic elements associated with resistance.

Isolation, host specificity and genetic characterization of Campylobacter specific bacteriophages from poultry and swine sources.

The isolation and characterization of 304 Campylobacter specific bacteriophage isolates from broiler and swine sources is reported in this study. Genome size characterization determined by PFGE classified these isolates,called CAM1-CAM304, within the campylophages group II (n = 18) and group III (n = 286). Host range analyses showed a high host specificity and similar lytic spectrum among isolates of the same group. Campylophages of group II infected C. jejuni, C. coli and even a C. fetus strain whereas those of group III only infected C. jejuni strains. The most promising 59 campylophage candidates were selected according to their lytic activity and their genetic diversity was analyzed by RFLP using SmiI and HhaI endonucleases for group II and III campylophages, respectively. Moreover, RAPD-PCR technique was for the first time assessed in the genetic characterization of campylophages and it was shown to be effective only for those of group II. Bacteriophage isolates grouped in a same genotype displayed different host ranges, therefore, 13 campylophages of group II and eight of group III were differentiated considering all the approaches assayed. An in-depth analysis of these bacteriophages will be performed to confirm their promising potential for the biocontrol of Campylobacter within the farm to fork process.

Evaluation of bacterial communities of Grana Padano cheese by DNA metabarcoding and DNA fingerprinting analysis.

The composition of the bacterial community of Grana Padano (GP) cheese was evaluated by an amplicon-based metagenomic approach (DNA metabarcoding) and RAPD-PCR fingerprinting. One hundred eighteen cheeses, which included 118 dairies located in the production area of GP, were collected. Two hundred fifty-four OTUs were detected, of which 82 were further discriminated between dominant (32 OTUs; > 1% total reads) and subdominant (50 OTUs; between 0.1% and 1% total reads) taxa. Lactobacillus (L.) delbrueckii, Lacticaseibacillus (Lact.) rhamnosus, Lact. casei, Limosilactobacillus fermentum, Lactococcus (Lc.) raffinolactis, L. helveticus, Streptococcus thermophilus, and Lc. lactis were the major dominant taxa ('core microbiota'). The origin of samples significantly impacted on both richness, evenness, and the relative abundance of bacterial species, with peculiar pattern distribution among the five GP production regions. A differential analysis allowed to find bacterial species significantly associated with specific region pairings. The analysis of pattern similarity among RAPD-PCR profiles highlighted the presence of a 'core' community banding pattern present in all the GP samples, which was strictly associated with the core microbiota highlighted by DNA metabarcoding. A trend to group samples according to the five production regions was also observed. This study widened our knowledge on the bacterial composition and ecology of Grana Padano cheese.

Molecular analysis of the dominant lactic acid bacteria of chickpea liquid starters and doughs and propagation of chickpea sourdoughs with selected Weissella confusa.

Fermented chickpea liquid is used as a leavening agent in chickpea bread production. In the present study, traditional chickpea liquid starter and dough samples were collected from bakeries in Turkey and microbiologically investigated. Culture-independent analysis for microbiota diversity, performed by MiSeq Illumina, identified Clostridium perfringens as major group in all samples, while Weissella spp. Dominated LAB community. A culture-dependent methodology was applied and 141 isolates were confirmed to be members of the LAB group based on 16s rRNA gene sequence analysis. In particular, 11 different LAB species were identified confirming the high frequency of isolation of weissellas, since Weissella confusa and Weissella cibaria constituted 47.8 and 12.4%, respectively, of total LAB isolated. The other species were Enterococcus faecium, Enterococcus lactis, Lactobacillus brevis, Lactobacillus plantarum, Leuconostoc mesenteroides, Leuconostoc mesenteroides subsp. Dextranium, Pediococcus acidilactici, Pediococcus pentosaceus and Streptococcus lutetiensis. Due to high frequency of isolation, W. confusa strains were investigated at technological level and W. confusa RL1139 was used as mono-culture starter in the experimental chickpea sourdough production. Chemical and microbiological properties, as well as volatile organic compounds (VOCs) of the chickpea liquid starters and doughs were subjected to a multivariate analysis. Control and W. confusa inoculated chickpea liquid starter and dough samples were close to each other in terms of some characteristics related to chemical, microbiological and VOCs profile, but the inoculated sourdough showed a higher generation of certain VOCs, like butanoic acid (81.52%) and ethyl acetate (8.15%) than control sourdough. This is important in order to maintain typical characteristics of the traditional chickpea dough, but at the same time improving the aroma profile. This work demonstrated that W. confusa RL1139 can be applied at large scale production level without compromising the typical characteristics of the final product.

Molecular characterization of Echinococcus granulosus in livestock of Al-Madinah (Saudi Arabia).

Echinococcus granulosus is the causative agent of cystic echinococcosis, which has serious impacts on human and/or animal health, resulting in significant economic losses. Echinococcus granulosus comprises a number of intra-specific variants or strains at the genetic level. In Saudi Arabia, few studies were performed on genetic variations in Echinococcus species. Therefore, the present study aimed to investigate the phenotypic and genetic characterization of hydatid cysts harboured by sheep and camels in Al-Madinah Al-Munawarah. Samples of hydatid cysts were collected from local sheep (n = 25) and camels (n = 8). The morphological criteria of protoscoleces were investigated. To investigate the molecular characterization, random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR), single-stranded conformation polymorphism (SSCP) were carried out. DNA was extracted from individual fertile cysts and subjected to RAPD-PCR analysis (using five arbitrary primers) and PCR amplification of cytochrome c oxidase I (cox1) and 12S ribosomal ribonucleic acid (12S rRNA) genes. The PCR products were subjected to SSCP analysis for genetic discrimination in E. granulosus isolates. In addition, partially sequencing of the mitochondrial DNA cox1 genes was achieved for assessing the phylogenetic positions of collected isolates using some global published sequence data of cox1 genes. The rostellar hooks of camel and local sheep isolates show remarkable variability in their dimensions. Five distinct SSCP patterns were identified in the 12S rRNA gene, showing intraspecific variations in E. granulosus of camels and local sheep. Sequencing of (cox1) genes of both local sheep and camels exhibit high similarity with those of the same gene (E. granulosus sensu stricto) published in NCBI BLAST.

Diversity and metallo-ß-lactamase-producing genes in Pseudomonas aeruginosa strains isolated from filters of household water treatment systems.

The microbiological quality of drinking water has long been a critical element in public health. Considering the high clinical relevance of Pseudomonas aeruginosa, we examined the filters of household water treatment systems for its presence and characteristics to determine the systems' efficiency in eliminating the bacteria. In total, filters of 50 household water treatment systems were examined. Microbiological and molecular methods were used for the detection and confirmation of P. aeruginosa isolates. Random Amplification of Polymorphic DNA-polymerase chain reaction (RAPD-PCR) was performed to detect similarities and differences among P. aeruginosa isolates. Combined disk (CD) method and double disk synergy test (DDST) were performed to detect metallo-beta-lactamase (MBL)-producing P. aeruginosa isolates. Finally, PCR was performed to detect MBL genes in MBL-producing strains. From the 50 analyzed systems, 76 colonies of P. aeruginosa were identified. In some systems, isolated bacteria from different filters harbored similar genetic profiles, indicating that these isolates may be able to pass through the filter and reach higher filters of the system. Phenotypic tests revealed 7 (9.2%) MBL-producing strains. Two isolates were positive for blaVIM-1, whereas one isolate was positive for blaNDM and blaIMP-1. The wide distribution of resistant phenotypes and genetic plasticity of these bacteria in household water treatment systems indicate that resistance mechanisms circulate among P. aeruginosa isolates in the environment of the filtration systems. The presence of MBL-producing genes in these systems and P. aeruginosa as a potential reservoir of these resistance genes can be a major concern for public health.

Growth performance, biochemical, cytological and molecular aspects of rabbits exposed to lead toxicity.

Heavy metals have enormous variety of deleterious effects on many organs in the body. This study demonstrated the toxic influences of lead on the growth, biochemical, cellular and molecular aspects of developing rabbits. Seventy-five rabbits (New Zealand NZW) were divided into five equal groups as follows; control (C) and four treatment groups (T1-4) orally administered lead acetate solution as follow T1: 20, T2: 30, T3: 50 and T4: 70 mg/kg body weight. Lead resulted in a significant decrease in live body weight, daily body weight gain and feed intake in T3 and T4 compared to those in other groups. Blood haematology measurements such as red blood cells, haematocrit (HCT), mean corpuscular volume, platelet, white blood cells and lymphocytes were significantly influenced by the high level of lead. Oral administration of lead significantly reduced total proteins in the serum. It was observed that the high lead level led to significantly (p < 0.05) increased levels of aspartate aminotransferase, alanine aminotransferase enzymes, urea and creatinine. Four random amplified polymorphic DNA primers polymorphism were detected among the treatment groups. Total number of induced bands (loss or appearance) compared with control group were 4, 10, 10 and 14 bands using primers P1, P2, P3 and P4 respectively. Number of micronuclei showed a dose-response increase and the difference was highly significant especially between control compared with T3 and T4 groups. From our results, we can conclude that exposure of rabbits to lead acetate resulted in negative effects on the growth performance and altered the haematological and biochemical parameters, in addition to its adverse impact on cytological and molecular characterization of animals.

Virulence factors, antibiotic resistance genes and genetic relatedness of commensal Escherichia coli isolates from dogs and their owners.

Escherichia coli (E. coli) is a normal flora of gastrointestinal tracts of humans and warm-blooded animals including dogs that has close vicinity with humans. Because the inter-species transmission of E. coli between pets and human beings, within a household, obtaining more information about the epidemiology, genetics, virulence factors, and antibiotic resistance of E. coli from dogs and their owners will help to control the inter-species transmission and treatment of E. coli infections. In this study we characterize and compare the antibiotic resistance and virulence profiles of fecal E. coli isolates from dogs and their owners. A total of 149 commensal E. coli isolates comprised 62 isolates from dogs, 56 isolates from their owners and 31 isolates from humans with no pet as control were collected. Extracted DNA was assessed for the presence of antibiotic resistance genes cmlA (chloramphenicol), sulI (sulfamethoxazole), floR (florfenicol) and blaCTX-M1 (cefotaxime) and virulence genes (papA, ompT, hlyD, traT, tsh and cnf1). To determine the extent of genetic relatedness of isolates, RAPD-PCR was performed. sulI and traT genes were the most dominant resistance profile and the most prevalent virulence gene in all groups, respectively, while hlyD had the lowest frequency among investigated virulence genes. Based on RAPD-PCR analysis clonal sharing between dogs and their owners were observed in 2/28 (7.1%) potential within-household clone-sharing pairs. Allowing dog to lick on owner's face, dog sex (female dogs), dog's sexual status (intact dogs) and times of disposing the feces (≥twice a day) were associated with a higher percentage of RAPD profile similarity (P < 0.05). The current study did not show an obvious evidence to prove considerable transmission of fecal E. coli from dogs to their owners. But in two households, there were relationship between isolates from dogs and their owners.

Serological and genetic characterization of Flavobacterium psychrophilum isolated from farmed salmonids in Turkey.

Turkey was the largest rainbow trout producer of the European countries in 2016, and the reason for this production is mainly attributed to its egg and fry production. Flavobacterium psychrophilum cause the highest rates of mortality in the starting to feeding stages of the fish. In the present study, twenty-five F. psychrophilum isolates recovered from rainbow trout, coruh trout and brook trout were analysed by RAPD-PCR, ERIC-PCR, REP-PCR and PCR-RFLP, including 16S rRNA, gyrA and gyrB gene regions and PCR-based serotyping method. The PCR-based molecular analysis showed that the isolates could not be differentiated exactly according to isolation source and geographical region. Most isolates were of type-1 and type-2, and some of them were of type-0 and type-3; in addition, one isolate showed a unique serotype. The combined analysis results showed that F. psychrophilum isolates discriminated as five different genotypes and all isolates were successfully discriminated based on host.

Production of probiotic bovine salami using Lactobacillus plantarum 299v as adjunct.

BACKGROUND: Five probiotic lactobacilli were tested, alone or in combination with two commercial starters, to select the most suitable strain for a probiotic bovine salami production. Lactobacillus plantarum 299v was used with both starters, to make salami according to a traditional recipe. Salami obtained by using just the starters and by spontaneous fermentation, served as control. Microbial dynamics, as well as the main physico-chemical parameters, were monitored throughout ripening. The survival of probiotic 299v was confirmed by strains' tracking by means of RAPD-PCR coupled to a culture-independent approach PCR-DGGE-based. RESULTS: The results showed a remarkable viability of the probiotic strain even after 60 days of storage. Experimental salami exhibited the same level of sensory acceptance of control salami, were hygienically safe, and characterised by pH, weight loss and microbiological loads within the ranges conventionally advocated for optimal fermented sausages. CONCLUSION: Outcomes indicate the workable possibility of using second-quality beef cuts for probiotic salami production. © 2017 Society of Chemical Industry.