Phenotypic and genotypic assessment of fluoroquinolones and aminoglycosides resistances in Pseudomonas aeruginosa collected from Minia hospitals, Egypt during COVID-19 pandemic.

BACKGROUND: One of the most prevalent bacteria that cause nosocomial infections is Pseudomonas aeruginosa. Fluoroquinolones (FQ) and aminoglycosides are vital antipseudomonal drugs, but resistance is increasingly prevalent. The study sought to investigate the diverse mechanisms underlying FQ and aminoglycoside resistance in various P. aeruginosa strains particularly during the COVID-19 crisis. METHODS: From various clinical and environmental samples, 110 P. aeruginosa isolates were identified and their susceptibility to several antibiotic classes was evaluated. Molecular techniques were used to track target gene mutations, the presence of genes encoding for quinolone resistance, modifying enzymes for aminoglycosides and resistance methyltransferase (RMT). Efflux pump role was assessed phenotypically and genotypically. Random amplified polymorphic DNA (RAPD) analysis was used to measure clonal diversity. RESULTS: QnrS was the most frequently encountered quinolone resistance gene (37.5%) followed by qnrA (31.2%) and qnrD (25%). Among aminoglycoside resistant isolates, 94.1% harbored modifying enzymes genes, while RMT genes were found in 55.9% of isolates. The aac(6')-Ib and rmtB were the most prevalent genes (79.4% and 32.3%, respectively). Most FQ resistant isolates overexpressed mexA (87.5%). RAPD fingerprinting showed 63.2% polymorphism. CONCLUSIONS: Aminoglycosides and FQ resistance observed in this study was attributed to several mechanisms with the potential for cross-contamination existence so, strict infection control practices are crucial.

Phytotoxicity and genotoxicity study of reactive red 141 dye on mung bean (Vigna radiata (L.) Wilczek) seedlings.

BACKGROUND: Reactive Red (RR) 141 dye is widely used in various industrial applications, but its environmental impact remains a growing concern. In this study, the phytotoxic and genotoxic effects of RR 141 dye on mung bean seedlings (Vigna radiata (L.) Wilczek) were investigated, serving as a model for potential harm to plant systems. METHODS AND RESULTS: Short-term (14 days) and long-term (60 days) experiments in paddy soil pot culture exposed mung bean seedlings to RR 141 dye. The dye delayed germination and hindered growth, significantly reducing germination percentage and seedling vigor index (SVI) at concentrations of 50 and 100 ml/L. In short-term exposure, plumule and radical lengths dose-dependently decreased, while long-term exposure affected plant length and grain weight, leaving pod-related parameters unaffected. To evaluate genotoxicity, high annealing temperature-random amplified polymorphic DNA (HAT-RAPD) analysis was employed with five RAPD primers having 58-75% GC content. It detected polymorphic band patterns, generating 116 bands (433 to 2857 bp) in plant leaves exposed to the dye. Polymorphisms indicated the appearance/disappearance of DNA bands in both concentrations, with decreased genomic template stability (GTS) values suggesting DNA damage and mutation. CONCLUSION: These findings demonstrate that RR 141 dye has a significant impact on genomic template stability (GTS) and exhibits phytotoxic and genotoxic responses in mung bean seedlings. This research underscores the potential of RR 141 dye to act as a harmful agent within plant model systems, highlighting the need for further assessment of its environmental implications.

Metal accumulation and genetic adaptation of Oryza sativa to Cadmiun and Chromium heavy metal stress: A hydroponic and RAPD analyses.

This research was performed to assess the influence of Cd and Cr metals on growth, pigments, antioxidant, and genomic stability of Oryza sativa indica and Oryza sativa japonica were investigated under hydroponic conditions. The results revealed that significant metal influence on test crop growth, pigment content, metal stress balancing antioxidant activity in a dose dependent manner. Since, while at elevated (500 ppm) concentration of Cd as well as Cr metals the pigment (total chlorophyll, chlorophyll a, b and carotenoids) level was reduced than control; however antioxidant activity (total antioxidant, H2O2, and NO) was considerably improved as protective mechanisms to combat the metal toxicity and support the plant growth. Furthermore, the test crops under typical hydroponic medium (loaded with Cd and Cr as 200, 300, 400, and 500 ppm) growth conditions, effectively absorb the metals from medium and accumulated in the root and least quantity was translocated to the shoot of this test crops. Furthermore, typical RAPD analysis with 10 universal primers demonstrated that the genomic DNA of the test crops was adaptable to develop metal resistance and ensure crop growth under increased concentrations (500 ppm) of tested heavy metals. These findings suggest that these edible crops have the ability to accumulate Cd along with Cr metals, and additionally that their genetic systems have the ability to adapt to metal-stressed environments.

Genetic relatedness and diversity of Capillaria species infecting bayad (Bagrus bajad) in upper Egypt.

BACKGROUND: This study investigates the genetic characteristics of Capillaria isolates from the infected fish, Bagrus bajad, and their relation to human Capillaria philippinensis using Random Amplified Polymorphic DNA (RAPD-PCR) analysis. Fifteen fish Capillaria were isolated and compared to identified human C. philippinensis using six primers: M-are, M-1, G-7, G-11, G-15, and G-18. RESULTS: All six primers successfully amplified DNA, highlighting their efficacy in distinguishing between human and fish Capillaria isolates. The analysis revealed distinctive banding patterns between fish and human isolates, with variations in size and number of DNA fragments. Additionally, genetic similarity analysis showed intriguing patterns of relatedness, with certain pairs exhibiting high similarity percentages. Comparative assessment of RAPD polymorphism demonstrated consistent findings of 100% polymorphism across all primers. The Unweighted Pair Group Method with Arithmetic Mean Algorithm (UPGMA) evaluated the closest relationship between human and fish isolates. These results underscore the utility of RAPD analysis in delineating the genetic diversity among Capillaria isolates from different hosts. CONCLUSION: Overall, this study contributes to our understanding of the genetic variability and relatedness among Capillaria isolates, shedding light on their evolutionary dynamics and zoonotic potential.

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.

Characterization of Coniella granati Isolates Causing Pomegranate Decline in Italy.

Coniella granati, the causal agent of pomegranate crown rot, twig blight, and fruit decay, is an emerging worldwide pathogen with a heavy impact on pomegranate cultivation. In this study, we report the rapid spread of the fungus in Italian pomegranate orchards associated with crown rot symptoms and provide new results on fungal development, baseline sensitivity to different fungicides, and intraspecific variability by analyzing 11 isolates, representative of populations of the pathogen from comparable pomegranate orchards in different regions of Italy. In vitro assays showed that 25 to 30°C was the optimal range for both colony growth and conidial germination, corroborating the results previously obtained for Californian and Greek isolates. According to the baseline sensitivity assay on the response of colony growth and conidial germination to 10 fungicides, fludioxonil, thiophanate-methyl, tebuconazole, and cyprodinil were the most effective. Random amplified polymorphic DNA (RAPD) analysis, carried out using fourteen 10-mer primers, showed very low intraspecific variability (similarity coefficient >0.95), probably as a result of the recent spread of the pathogen in Italy and the uncommon occurrence of the sexual process as a source of genetic variability. In summary, this study provides new knowledge on C. granati that will be helpful for improving pomegranate crown rot management.

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.

Genotyping-driven diversity assessment of biocontrol potent Bacillus spp. strain collection as a potential method for the development of strain-specific biomarkers.

Bacillus species are among the most researched and frequently applied biocontrol agents. To estimate their potential as environmentally friendly microbial-based products, reliable and rapid plant colonization monitoring methods are essential. We evaluated repetitive element-based (rep) and Random Amplified Polymorphic DNA (RAPD) PCR (Polymerase Chain Reaction) genotyping in a diversity assessment of 251 strains from bulk soil, straw, and manure samples across Serbia, highlighting their discriminative force and the presence of unique bands. RAPD 272, OPG 5, and (GTG)5 primers were most potent in revealing the high diversity of a sizable environmental Bacillus spp. collection. RAPD 272 also amplified a unique band for a proven biocontrol strain, opening the possibility of Sequence Characterized Amplified Region (SCAR) marker design. That will enable colonization studies using the SCAR marker for its specific detection. This study provides a guide for primer selection for diversity and monitoring studies of environmental Bacillus spp. isolates.

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.

Evaluating genetic diversity of geographically diverse populations of Embelia ribes Burm f., a highly medicinal woody liana from the Western Ghats of India, using random amplified polymorphic DNA (RAPD) and intersimple sequence repeats (ISSR) markers.

BACKGROUND: Embelia ribes Burm f. (Primulaceae) is a medicinal and vulnerable woody liana distributed throughout India. Embelin, a well-recognized active phytoconstituents in berries, is commonly used in ayurvedic formulations. Due to over-exploitation, the status of the plant is vulnerable. Previous studies on this species mainly focused on its phytochemical analysis, which led to overexploitation and loss of the germplasm. METHODS AND RESULTS: In the present study, 20 RAPD and 18 ISSR markers were employed to assess genetic divergence in 40 genotypes of E. ribes collected from different parts of the Western Ghats of India. In RAPD analysis, all 40 accessions with 20 RAPD primers amplified 282 fragments, with 83.91% average polymorphism and with an average of 14.10 bands per primer. The size of amplicons varied from 200 to 2500 bp. While, ISSR primers produced 203 fragments of which 161 were polymorphic with an average of 11.28 bands per primer with 73.25% average polymorphism. The size of amplicons ranges from 200 to 2500 bp. RAPD and ISSR markers were also assessed by calculating polymorphic information content (PIC) to discriminate the genotypes; the average PIC value for RAPD, ISSR, and combined RAPD + ISSR markers obtained was more than 0.50 suggesting the informativeness of markers. UPGMA analysis based on Jaccard's similarity coefficient for RAPD, ISSR, and RAPD + ISSR data reveals that 40 accessions of E. ribes were depicted in four clusters. The clustering pattern of all individuals in PCoA analysis agreed with the UPGMA dendrograms, which further confirms the genetic relationships explained by cluster analysis. AMOVA analysis of RAPD, ISSR, and combined marker system revealed variation within the population, ranging from 41 to 44%, and among the population, it ranged from 56 to 59%. CONCLUSION: The present study provides an optimized method for evaluating the genetic diversity of Embelia ribes using RAPD and ISSR markers which are useful for further sustainable utilization and conservation of natural populations in the Western Ghats of India.

Mitotic chromosomal abnormalities and DNA polymorphism in the Nile tilapia (Oreochromis niloticus, Linnaeus, 1758) as a biomarker for water pollution by heavy metals.

Mitotic chromosomal aberrations and DNA polymorphism (RAPD marker) were carried out on the Nile tilapia Oreochromis niloticus collected from five sites in Minia governorate, Egypt to test their applicability as biomonitors for heavy metal contaminants of water. The diploid chromosome number of O. niloticus population was 2 n = 44. Different types of chromosomal aberrations were recorded (e.g., deletion, ring, centromeric attenuation, end-to-end association, dicentric chromosome, stickiness chromosomes, endomitosis, fragments and chromatid gap). The chromosomal aberrations varied between O. niloticus population collected from five sites, and the most common type was ring (R) chromosomes. Samples obtained from Bahr Yousef and Irrigation drain exhibited the highest aberration frequency. The frequency of chromosomal aberration was positively correlated with the concentration of heavy metals where their concentration in the surface water of Irrigation drain and Bahr Yousef exceeded the limits defined by WHO as well as the concentration of Pb in muscles. The RAPD marker was also used to identify genetic variation among Nile tilapia samples collected from five different water sources. It created polymorphic and unique bands that can be used as genetic markers to track DNA variations. The dendrogram also revealed that exposure to heavy metal pollution causes gradual accumulation of variance, whereas areas subjected to environmental stress showed higher genetic variation and clustered together.

Rindera graeca (A. DC.) Boiss. & Heldr. (Boraginaceae) In Vitro Cultures Targeting Lithospermic Acid B and Rosmarinic Acid Production.

The in vitro cultures of Rindera graeca, a rare endemic plant, were developed as a sustainable source of phenolic acids. Various shoot and root cultures were established and scaled up in a sprinkle bioreactor. A multiplication rate of 7.2 shoots per explant was achieved. HPLC-PDA-ESI-HRMS analysis revealed the presence of rosmarinic acid (RA) and lithospermic acid B (LAB) as the main secondary metabolites in both the shoot and root cultures. The maximum RA (30.0 ± 3.2 mg/g DW) and LAB (49.3 ± 15.5 mg/g DW) yields were determined in root-regenerated shoots. The strongest free radical scavenging activity (87.4 ± 1.1%), according to 2,2-diphenyl-1-picrylhydrazyl-hydrate assay, was noted for roots cultivated in a DCR medium. The highest reducing power (2.3 µM ± 0.4 TE/g DW), determined by the ferric-reducing antioxidant power assay, was noted for shoots cultivated on an SH medium containing 0.5 mg/L 6-benzylaminopurine. A genetic analysis performed using random amplified polymorphic DNA and start codon targeted markers revealed genetic variation of 62.8% to 96.5% among the investigated shoots and roots. This variability reflects the capacity of cultivated shoots and roots to produce phenolic compounds.

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.

Molecular and metabolic traits of some Egyptian species of Cassia L. and Senna Mill (Fabaceae-Caesalpinioideae).

The genus Cassia and Senna have been classified under subfamily Caesalpinioideae of family Fabaceae (Leguminosae) of order Fabales. There is a scarce taxonomical studies of the genus Cassia and Senna inhabiting Egyptian environments, thus, the main objective of the current was to revise and authenticate the phylogenetic relationship between studied taxa of the species of the genera Cassia and Senna in Egypt using the recent tools of ITS barcoding, RAPD analysis and metabolic profiling, in comparing to the traditional taxonomical features. From the cluster analysis of the traditional 27 morphological characters, the studied taxa were categorized into two major clades with an average taxonomic distance of 4.3. The clade I include Cassia fistula, C. renigera, C. javanica L subsp. nodosa and C. roughiia that belongs to series Obolospermae, and C. grandis that belongs to series Grandes. The clade (II) includes Senna surattensis and S. alata at taxonomic level 3.6. The taxonomical description of the studied taxa was confirmed from the molecular analysis of ITS sequences and RAPD analysis. The ITS sequences of the tested plants species C. fistula L, C. grandis MD4, C. javanica subsp. nodosa MD7, C. roxburghii MD5, C. renigera MD5 were deposited at genbank with accession numbers MW367973, MZ960447, MW386305, MW326753 and MW32685, respectively. While, the ITS sequences of the S. surrattensis and S. alata were deposited into genbank accession # MD14 MW367670 and MD20 MW412635, respectively. Thus, from the molecular analysis, two clades were clearly separated into Clade I of Cassia and Clade II of Senna. The cluster I represented by C. fistula, C. renigera, C. roxburghii, and C. javanica sub nodosa, and the cluster II represented by S. alata and S. surattensis. From the PCA of RAPD, a clearly discrimination between the two Taxa was observed revealing the characteristic grouping of Cassia and Senna. The species Senna alata and Senna surattensis were grouped together, but the species of C. renigera, C. javanica, C. roxburghii and C. grandis was grouped on a distinct group. The separation of Cassia and Senna species into two clusters verify the segregation of the genus Cassia L. senso lato into two distinct genera namely Senna P. and Cassia L. The morphological, molecular traits of the studied plants were authenticated from the metabolic profiling by GC-MS analysis. Among the 23 identified metabolites, four compounds namely hexadecanoic acid, methyl ester, 9-Octadecenoic acid (Z)-ethyl ester and Vitamin E were detected with fluctuated concentrations, among C. fistula, C. grandis, C. javanica subsp. nodosa and C. roxburghii. Conclusively, the traditional morphological features, molecular barcoding using ITS sequences, RAPD analysis and metabolic traits by GC-MS analysis, authenticates the taxonomical diversity of the genus Cassia and Senna.

Elucidating genetic diversity and population structure of Pyricularia oryzae isolates causing wheat blast in Bangladesh.

Wheat blast, caused by Pyricularia oryzae pathotype Triticum, is one of the most notorious diseases of wheat. In this study, a total of twenty-four monoconidial isolates representing four major wheat blast affected districts, namely Chuadanga, Meherpur, Kustia and Jhenaidah of Bangladesh were analyzed. Eight RAPD and four ISSR primers being used for genetic diversity assay produced a total of 94 bands of which 85% were polymorphic. UPGMA dendrogram based on combined dataset (RAPD and ISSR) separated all the isolates into two main clusters having similarity ranged from 64 to 93%. Principal coordinate analysis showed congruent result with cluster analysis. However, clustering of the isolates according to their origin and plant part sampled was not apparent in the dendrogram. The genetic diversity indices unveiled that genetic diversity in P. oryzae populations is low. Average Nei's gene diversity (h) and Shannon's Information Index (I) calculated for isolates from each district were found 0.16 and 0.24, respectively. The population structure analysis of the isolates revealed the presence of two sub-populations with admixture of alleles. Analysis of molecular variance indicated that significantly higher level of variation (96%) in the population was present within districts while a relatively low proportion (4%) of the variation was detected among districts. Knowledge generated in this study will give a pace in the development of appropriate wheat blast management strategies to control this disease in Bangladesh.

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.

Diversity of endophytic bacteria in hybrid maize seeds and Bacillus mojavensis J2416-7 may be capable of vertical transmission.

The diversity of endophytic bacteria in the progeny is related to the parental lines. In this study, the traditional separation method was used to study the dominant endophytic bacteria of the shared paternal line and its pollen, different maternal lines and their F1 progeny. And the results showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus and Pantoea. The Bacillus diversity of the progeny JMC121 and JN728 were the same as both the paternal line and the maternal line, including Bacillus subtilis, Bacillus velezensis, Bacillus mojavensis, and Bacillus licheniformis. The Bacillus subtilis and Bacillus velezensis in JN828 were the same as both the paternal line and the maternal line, while Bacillus licheniformis was only the same as the paternal line. Through the RAPD molecular typing, there was the same strain of Bacillus mojavensis existed in the paternal line J2416, the pollen and the progeny JN728; this meant that the paternal line passed its dominant endophytic bacteria to the progeny through pollen in vertical transmission. This study showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus, and the diversity of F1 progeny was related to both the paternal line and the maternal line.

Comparative Analysis Between Paramecium Strains with Different Syngens Using the RAPD Method.

Paramecium spp. are a genus of free-living protists that live mainly in freshwater environments. They are ciliates with high motility and phagocytosis and have been used to analyze cell motility and as a host model for pathogens. Besides such biological characteristics, apart from the usual morphological and genetic classification of species, the existence of taxonomies (such as syngens) and mating types related to Paramecium's unique reproduction is known. In this study, we attempted to develop a simple method to identify Paramecium strains, which are difficult to distinguish morphologically, using random amplified polymorphic DNA (RAPD) analysis. Consequently, we can observe strain-specific band patterns. We also confirm that the presence of endosymbiotic Chlorella cells affects the band pattern of P. bursaria. Furthermore, the results of the RAPD analysis using several P. caudatum strains with different syngens show that it is possible to detect a band specific to a certain syngen. By improving the reaction conditions and random primers, based on the results of this study, RAPD analysis can be applied to the identification of Paramecium strains and their syngen confirmation tests.

Development of molecular markers linked to QTL/genes controlling Zn efficiency.

BACKGROUND: Zinc (Zn) deficiency is a widespread problem in reducing the yield and quality of crop plants worldwide. It is important to utilize molecular markers linked to Zn efficiency to develop high Zn efficient cultivars in pepper (Capsicum annuum L.). METHODS AND RESULTS: In present study, genetic map was constructed using a F2 populations derived from C. annuum L. (Alata 21A) × C. frutescens L. (PI 281420) cross. The quantitative trait locus (QTLs) for Zn efficiency were mapped using F2:3 population. A genetic map with 929.6 cM in length and 12 linkage groups were obtained using 62 markers (31 sequence-related amplified polymorphism (SRAP), 19 simple sequence repeat (SSR) and 11 random amplified polymorphic DNA (RAPD) markers). The 41 linked QTLs related with nine (9) Zn efficiency characters were mapped on linkage groups. Results suggest that selecting plants for tolerance to Zn deficiency are expected to be rather responsive among segregating populations for breeding and developing Zn efficient genotypes in pepper. CONCLUSIONS: The molecular markers are expected to aid selection and expedite breeding peppers resistant to Zn deficiency in soils low for available Zn contents.

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.