An investigation of the genotoxic and cytotoxic effects of myclobutanil fungicide on plants

Myclobutanil is a chemical pesticide commonly used in the production of some vegetables and fruits like greenhouse peppers, grapes, and apples. The aim of this study was to investigate the genotoxic and cytotoxic effects of myclobutanil fungicide on the Allium cepa plant, the model organism. Randomly amplified polymorphic DNA and inter simple sequence repeat-PCR techniques were performed on the DNA of A. cepa exposed to the different myclobutanil doses and time periods. The nucleus anomalies and abnormal anaphases were investigated using a light microscope. PCR analyses showed that myclobutanil causes some DNA sequence changes on the onion genome depending on the increase in the fungicide dose and exposure time. It was determined that myclobutanil has a serious genotoxic effect, even in low doses like 25–50 ppm.

ANALYSIS OF GENETIC DIVERSITY OF 26 CASSAVA VARIETIES (Manihot esculenta CRANTZ) WITH DIFFERENT RESPONSES TO WATERLOGGING CONDITIONS BY USING RAPD MARKERS

RAPD (Randomly Amplified Polymorphic DNA) is an indicator for high and stable polymorphism, widely used in the study of the diversity of cassava. In this paper, the results of using 20 polymorphic primers OPK combined with the establishment of the phylogenetic tree to analyze the genetic diversity of 26 cassava varieties with different responses to waterlogging conditions by using the RAPD-PCR technique were presented. The purpose of this experiment was to show the genetic relevance of the studied cassava varieties. The results showed that the flood tolerance of cassava was not related to the polymorphism and branching characteristics of the stem. This information may be use as a basis for selecting flood-tolerant cassava varieties for cassava production, as well as the basis for selecting genetically different parents for breeding.

Investigations on Transfer of Pathogens between Foster Cows and Calves during the Suckling Period

To date, there have been few studies on the health effects of foster cow systems, including the transmission of mastitis-associated pathogens during suckling. The present study aimed to compare the pathogens detected in the mammary glands of the foster cow with those in the oral cavities of the associated foster calves and to evaluate the resulting consequences for udder health, calf health and internal biosecurity. Quarter milk sampling of 99 foster cows from an organic dairy farm was conducted twice during the foster period. Oral cavity swabs were taken from 345 foster calves. Furthermore, quarter milk samples were collected from 124 biological dams to investigate possible transmission to the foster cows via the suckling calves. All samples were microbiologically examined and confirmed by MALDI-TOF (matrix-assisted laser desorption time-of-flight mass-spectrometry). Using RAPD-PCR (randomly amplified polymorphic DNA polymerase chain reaction), strain similarities were detected for Pasteurella multocida, Staphylococcus aureus, S. sciuri and Streptococcus (Sc.) suis. Transmission of P. multocida and S. aureus probably occurred during suckling. For S. sciuri and Sc. suis, environmental origins were assumed. Transmission from dam to foster cow with the suckling calf as vector could not be clearly demonstrated.

Degenerate primed randomly amplified polymorphic DNA (DP-RAPD) fingerprinting of bacteriophages of Vibrio harveyi from shrimp hatcheries in Southern India

Vibrio harveyi is a significant pathogen of shrimp. Seventy-six bacteriophages infecting luminescent V. harveyi were isolated from a total of 194 water samples drawn from various sources of shrimp hatcheries located in South East coast and Andaman island of India. Degenerate primed randomly amplified polymorphic DNA (DP- RAPD) fingerprinting of these bacteriophages was carried out to determine their genetic relatedness. Similarity matrix based on Dice coefficient followed by construction of dendrogram by unweighted pair group method with arithmetic averages (UPGMA) revealed 12 major clusters. One phage was randomly selected from each of these major clusters for transmission electron microscopic observations. Eleven of them had an icosahedral head (46-115 nm) with a long non- contractile tail (132-329 nm), belonging to the Siphoviridae family and two phages had a short tail (15-27 nm), belonging to the family Podoviridae. The phylogenetic analysis of the phages using DP-RAPD fingerprinting correlated to some extent to the phenotypic nature of the host specifically with regard to sucrose fermentation and source of isolation. However, phages specifically infecting V. harveyi and those belonging to different families did not cluster together in the DP-PCR cluster analysis. Hence, the genetic diversity of phages infecting same host with respect to phenotypic difference was revealed by the DP-RAPD applied in this study.

VARIASI GENETIK TIGA GENERASI IKAN HIAS CUPANG ALAM ENDEMIK DARI ACEH Betta rubra, Perugia 1893 (Pisces: Osphronemidae), HASIL BUDIDAYA

Betta rubra merupakan salah satu spesies ikan cupang alam endemik dari Aceh. Keberadaannya yang hampir dinyatakan punah sebelum ditemukan kembali pada tahun 2007. Tujuan dari penelitian ini adalah mengkaji keragaman genetik dan potensi genetik dari ikan Betta rubra dari tiga generasi yang sudah dibudidayakan untuk perbaikan genetik di Balai Riset Budidaya Ikan Hias (BRBIH), Depok, Jawa Barat, Indonesia. Jumlah sampel yang digunakan pada populasi G-0 adalah enam ekor, sedangkan pada populasi G-1 dan G-2 masing-masing 10 ekor. Ikan uji yang digunakan diambil sirip ekornya untuk analisis secara genotipe dengan randomly amplified polymorphic DNA (RAPD) menggunakan primer yaitu OPZ-9, OPB-6, dan OPZ-13. Sebelum diambil sirip ekornya ikan terlebih dahulu difoto di atas millimeter block untuk data truss morfometrik (fenotipe). Hasil menunjukkan ikan Betta rubra populasi alam (G-0) memiliki nilai heterozigositas 0,1872 dan derajat polimorfisme 47,06% yang lebih rendah dibandingkan generasi G-1 dengan heterozigositas 2,421 dan derajat polimorfisme 64,71%. Populasi G-2 memiliki nilai heterozigositas 0,1577 dan derajat polimorfisme 44,12%. Koefisien keragaman secara fenotipe populasi G-1 memiliki variasi lebih tinggi dibanding populasi G-0 dan G-2. Hubungan kekerabatan antara G-1 dengan G-0 dan G-2 berbeda nyata (P<0,05), sedangkan hubungan antara G-1 dengan G-2 tidak berbeda nyata (P>0,05), sehingga antara populasi G-0 dan G-2 membentuk cluster terpisah dengan G-1. Keragaman genetik pada tiga generasi Betta rubra memiliki pola yang sama baik secara fenotipe maupun genotipe.Betta rubra is one of the endemic species of Betta fish from Aceh. The fish was almost declared extinct before it was rediscovered in 2007. The purpose of this study was to examine the genetic diversity and genetic potential of Betta rubra from three generations which have been reared for genetic improvement at the Research Institute for Ornamental Fish Culture, Depok, West Java, Indonesia. The number of fish for G-0 population used in the study was six fish whilst G-1 and G-2 populations were 10 fish. Tail fins from each fish were sampled for genotype analysis using randomly amplified polymorphic DNA (RAPD) using primers OPZ-9, OPB-6, and OPZ-13. Before tail fin collection, the fish was photographed on a millimeter block for truss morphometric data measurement (phenotype). The results showed that the Betta rubra wild population (G-0) had heterozygosity of 0.1872 and polymorphism of 47.06% which were lower than the G-1 population with heterozygosity of 2.421 and polymorphism of 64.71%. The G-2 population had heterozygosity of 0.1577 and polymorphism of 44.12%. The phenotype coefficient of variation in the G-1 population higher than the G-0 and G-2 populations. The kinship relationship between G-1 with G-0 and G-2 was significantly different (P<0.05), while the relationship between G-1 and G-2 was not significantly different (P>0.05). This research concludes that the populations of G-0 and G-2 have formed a separate cluster to G-1. The genetic diversities in the three Betta rubra populations have similar phenotype and genotype patterns.

Study of arsenic genotoxicity in a freshwater fish (Channa punctatus) using RAPD as molecular marker

Arsenic, the proven genotoxic carcinogen in humans, is a groundwater contaminant of global concern. The present work investigates the applicability of randomly amplified polymorphic DNA (RAPD) as molecular marker to demonstrate arsenic genotoxicity in the freshwater fish, Channa punctatus. Fish specimens, segregated into three groups, were exposed to 10, 50 and 500 µgL− 1 of arsenic respectively for 20 consecutive days. DNA extracted from same specimens of each group before and after exposure was amplified by polymerase chain reaction (PCR) using single arbitrary primers. Marked changes in RAPD profiles of fish DNA were observed upon exposure to arsenic compare to RAPD profiles of their pre-exposure state, resulting from loss or gain of certain bands. Altogether a total of 41 loci were amplified with 37–41 bands in each group. Band 1 of pre-exposure state was lost in all post-exposure samples while bands 4, 6 and 7 appeared as new bands after exposure to arsenic. The changes in the RAPD banding patterns upon exposure to arsenic reflect alterations in fish DNA. The RAPD bands, therefore, appeared as potential markers, capable of revealing the genotoxicity induced by arsenic in this piscine model.

Randomly amplified polymorphic DNA analysis of Staphylococcus chromogenes isolated from bovine and bubaline mastitis in Karnataka

Background and Aim: In recent times, non-aureus staphylococci (NAS) have emerged as the major organisms isolated from mastitis cases in dairy animals, with a predominance of Staphylococcus epidermidis and Staphylococcus chromogenes. As compared to Staphylococcus aureus, much less is known about the molecular types or the spatiotemporal epidemiology of these NAS species. In the present study, randomly amplified polymorphic DNA (RAPD) was employed to detect genetic polymorphisms, intraspecies diversity, and epidemiology of S. chromogenes strains (n=37) isolated from bovine and bubaline mastitis cases in the state of Karnataka. Materials and Methods: Thirty-seven S. chromogenes isolates (14 from bovines and 23 from bubaline) isolated from subclinical mastitis cases, from organized and unorganized sectors, were subjected to RAPD typing. Further, methicillin resistance was determined by cefoxitin disk diffusion method. Results: The amplified DNA fragments ranged from 150 to 3000 base pairs and yielded several RAPD profiles. Further analysis using Digital Image Correlation Engine correlation coefficient and UPGMA method showed that the 37 isolates could be classified into 12 distinct RAPD types (A to L) at 62% similarity (D=0.889). Four of the most predominant RAPD types, B, A, C, and E, in that order, and together, represented 65% of the isolates. High diversity was observed among the isolates both within farms and between geographic locations. Most of the isolates exhibited methicillin resistance. This is the first such report from India. Conclusion: In the absence of defined multilocus sequence type protocols or sufficient sequences available in the public domain, RAPD can be employed to determine genetic diversity of S. chromogenes isolates.

Genetic Similarity of Commerson’s Anchovy across Segara Anakan Cilacap Assessed Using Randomly Amplified Polymorphic DNA (RAPD) Markers

Segara Anakan areas can be divided into three different regions according to their salinity. Salinity differences suggested that Commerson’s anchovy population in that area can be divided into three subpopulations due to genetic differences. Genetic differences among subpopulation can be assessed through a population genetic study using random amplified polymorphic DNA. This study aims to evaluate the genetic variation and differences of Commerson's anchovy (Stolephorus commersonnii) collected at three different water salinities in Segara Anakan estuary Cilacap Indonesia. Total genomic DNA was isolated using the Chelex method. Genetic diversity and differences were assessed using RAPD markers and were analyzed statistically using an analysis of molecular variance, as implemented in Arlequin software. The results showed that high genetic diversity was observed within the subpopulations. However, no significant genetic differences were observed among subpopulations which indicate genetic similarity. A high number of offspring are likely to cause high genetic variation within subpopulations. Adult and larvae migration is the cause of genetics similarity across Segara Anakan. Another impressive result is that water salinity did not affect the genetic characteristic of Commerson,s anchovy. Genetic similarity of Commerson’s anchovy indicates that Segara Anakan forms a single genetic conservation unit.