Polymorphic DNA Research Articles

High Frequency of blaOXA-48like Producing Klebsiella pneumoniae Isolated from Nosocomial Infection in Azerbaijan, Iran.

Klebsiella pneumoniae is one of the significant agents of hospital-acquired infections. In recent years, carbapenem-resistant K. pneumoniae (CRKP) isolates have been found in numerous epidemics of nosocomial infections. This study aimed to determine carbapenem resistance mechanisms and molecular epidemiological of CRKP infections in Azerbaijan, Iran. A total of 50 non-duplicated CRKP from January 2020 to December 2020 were isolated form Sina and Imam Reza Hospitals in Tabriz, Iran. Antimicrobial susceptibility testing was performed by the disk-diffusion method. The carbapenem resistance mechanisms were determined by the phenotypic and PCR procedures. CRKP isolates were typed by the Random Amplified Polymorphic DNA PCR (RAPD-PCR) technique. Amikacin was the most effective antibiotics against CRKP isolates. AmpC overproduction was observed in five CRKP isolates. Efflux pump activity was found in one isolate by the phenotypic method. Carba NP test could find carbapenemases genes in 96% of isolates. The most common carbapenemases gene in CRKP isolates were blaOXA-48-like (76%) followed by blaNDM (50%), blaIMP (22%), blaVIM (10%), and blaKPC (10%). The outer membrane protein genes (OmpK36 and OmpK35) were identified in 76% and 82% of CRKP isolates, respectively. RAPD-PCR analysis yielded 37 distinct RAPD-types. Most blaOXA-48-like positive CRKP isolates were obtained from patients hospitalized in intensive care unit (ICU) wards with urinary tract infections. The blaOXA-48-like is the main carbapenemase among CRKP isolates in this area. Most blaOXA-48-like producer CRKP strains were collected from the ICU ward and urine samples. To control infections due to CRKP, a strict control program in hospital settings is required.

Pseudomonas Aeruginosa Colonising Patients with Cystic Fibrosis: Antibiotic Resistance and Growth Conditions Determined by in vivo and in vitro Gene Expression

Background: Pseudomonas aeruginosa is the main pathogen responsible for lung destruction in cystic fibrosis, becoming difficult to eradicate in chronic infection. Aims: To describe antibiotic resistance among strains of P.aeruginosa isolated from sputa of patients with cystic fibrosis. To investigate in vivo and in vitro expression of genes related to antibiotic resistance and anaerobic growth. Methods: Sputa (in vivo) and strains (in vitro) from 26 patients were obtained during 17 months. Genotypes were compared by random polymorphic DNA amplification. Expression of nirS (anaerobic respiration) and mexY (MexXY efflux pump) were measured by quantitative real time polymerase chain reaction. Expression levels of nirS in aerobiosis and anaerobiosis were compared to estimate oxygenation status within lungs. Mutations in the regulator gene mexZ were investigated in sputa expressing mexY and were correlated with strains’ antibiotic resistance. Results: Nine patients and 56 sputa were finally analysed. Seven patients carried a single genotype. Gene mexY was detected in all the sputa; expression levels were higher in sputa with mexZ mutations. Multi-resistance was frequent. Resistance profiles not always correlated with mexY expression levels or mexZ mutations. Comparison of in vivo and in vitro nirS expression indicated mainly aerobic and microaerophilic environments within sputa. Discussion: Mutations in e mexZ are frequent in strains of P.aeruginosa colonising patients with cystic fibrosis. Presence of these mutations correlates with increased expression of mexY in vivo and in vitro, but no with in vivo antibiotic resistance. Results of nirS expression suggest that the lungs represent heterogeneous environments regarding oxygenation status. This complexity explains that mechanisms of growth and antibiotic resistance within the lungs of these patients are still largely unknown. Conclusions: After many years of research few studies, including the present, revealed different aspects of in vivo growth of P. aeruginosa. We determined a cut-off to discriminate between sputa containing mexZ wild type and mutated alleles and showed that comparison of in vivo and in vitro nirS expression allows to predict oxygenation status. So far, none of the studies can explain all the factors influencing the behaviour of P.aeruginosa colonising cystic fibrosis patients making it difficult to design new therapeutic strategies.

Characterization and variability of strains of Xanthomonas axonopodis pv. passiflorae from the state of Pará, Brazil

Bacterial spot, caused by Xanthomonas axonopodis pv. passiflorae, is a disease that has limited the cultivation of passionfruit in various orchards in Brazil. The objective of this work is to characterize and evaluate the variability of 29 strains of X. axonopodis pv. passiflorae from different municipalities producing yellow passionfruit in the state of Pará. The characterization was performed by the biochemical methods of KOH solubility, oxidase and Bactray, and the molecular methods of Xapas-F, Xapas-Ri and Xapas-Ro primers. The variability was evaluated by pathogenicity test and RAPD (randomly amplified polymorphic DNA). The strains of X. axonopodis pv. passiflorae were Gram-negative and oxidase-negative, and tests with the Bactray kit showed no relation between the collection municipality and group composition. The Xapas-F/Ri/Ro primers were specific for the strains. The primers used amplified 118 polymorphic bands in the RAPD reactions and the highest genetic similarity was between the strains PA15 and PA16. As the pathogenicity test evidenced a pathogenic variability, the strains PA2.1, PA4.5, PA14, PA4.2, PA4.1, PA4.6, PA3.4 and PA4.3 present the highest severity values for the disease. The strains of X. axonopodis pv. passiflorae show characteristics typical of the species, and genetic and pathogenic variability among them.

Random Amplified Polymorphic DNA for identical Streptococcus salivarius strains isolated from tongue of peoples before and after Listerine In vivo

Random Amplified Polymorphic DNA for identical Streptococcus salivarius strains isolated from tongue of peoples before and after Listerine In vivo

Induction of genetic variability with gamma radiation and detection of DNA polymorphisms among radiomutants using sequence-related amplified polymorphism markers in Gaillardia pulchella Foug. plants

Induction of genetic variability with gamma radiation and detection of DNA polymorphisms among radiomutants using sequence-related amplified polymorphism markers in Gaillardia pulchella Foug. plants

V-primer: software for the efficient design of genome-wide InDel and SNP markers from multi-sample variant call format (VCF) genotyping data.

DNA markers are indispensable tools in genetics and genomics research as well as in crop breeding, particularly for marker-assisted selection. Recent advances in next-generation sequencing technology have made it easier to obtain genome sequences for various crop species, enabling the large-scale identification of DNA polymorphisms among varieties, which in turn has made DNA marker design more accessible. However, existing primer design software is not suitable for designing many types of genome-wide DNA markers from next-generation sequencing data. Here, we describe the development of V-primer, high-throughput software for designing insertion/deletion, cleaved amplified polymorphic sequence, and single-nucleotide polymorphism (SNP) markers. We validated the applicability of these markers in different crops. In addition, we performed multiplex PCR targeted amplicon sequencing using SNP markers designed with V-primer. Our results demonstrate that V-primer facilitates the efficient and accurate design of primers and is thus a useful tool for genetics, genomics, and crop breeding. V-primer is freely available at https://github.com/ncod3/vprimer.

Chloroplast genomes of Vitis sp. samples from Nechaevsk population of the Krasnodar region

The presented study demonstrates the results of sequencing and analysis of wild grape plastomes assemblies located in the Nechaevsk population. Previously, the habitats of the wild forest grape V. vinifera subsp. silvestris were explored as an outcome of expeditions. As a result of repeated expeditions, samples were taken and used to isolate chloroplast DNA. Assembly and annotation of plastomes showed the presence of DNA encoding sequences. At the same time, the structure of the obtained plastomes varied. These expressed in the presence of DNA polymorphisms. Comparison of the aligned sequences made it possible to reveal the presence of the species V. vinifera subsp. sylvestris in the population, as well as hybrid specimens. This is also confirmed by the result of determining the taxonomy after depositing the archives of readings in the NCBI database.

Генетическое разнообразие российских и европейских пред-ставителей Dryopteris expansa (C. Presl) Fraser-Jenkins et Jermy (Dryopteridaceae)

The results of molecular genetic analysis (ISSR-PCR) of 112 Dryopteris expansa (C. Presl) Fraser-Jenkins et Jermy specimens collected in Russia and Europe are presented. DNA was extracted using fresh material and material from the herbarium of the South Siberian Botanical Garden (ALTB): the collection of fresh material was carried out during expeditions (40 samples from Perm Krai, Moscow, Smolensk and Tver regions), the selection of material from the herbarium was carried out by removing a part of the dried frond (72 samples from various regions of Russia and Europe). An analysis of DNA polymorphism and the genetic structure of the studied groups with the calculation of the proportion (P95) of polymorphic loci, the absolute (na) number of alleles, the effective (ne) number of alleles, the expected (HE) heterozygosity and the information-entropy measure (I) of Shannon was carried out using the POPGENE 1.31 program. To describe the genetic structure of the studied groups, the following parameters were used: the expected proportion of heterozygous genotypes (HT) in the entire group as a measure of its overall diversity; the expected proportion of heterozygous genotypes in a subgroup (HS) as a measure of its internal diversity; the proportion of intergroup genetic diversity in the total diversity or the subdivision index (GST). In addition to this, the AMOVA package was used with the calculation of the subdivision index of groups using 1000 rounds of permutations. Based on a comparison of the indicators of the Shannon information measure (I), Nei fixation index (FST), as well as intra- and intergroup variability based on the results of the analysis of molecular variants (AMOVA) obtained for D. expansa with similar indicators given for D. fragrans (L.) Schott, a conclusion is made about the inexpediency of dividing D. expansa s.l. into two independent species – D. expansa s. str. and D. assimilis S. Walker. The indicators of all indices and calculated values given in the publication can be used as a basis for comparison in other works devoted to the genetic variability of species of the genus Dryopteris in Russia.

Angiotensinogen, Angiotensin-Converting Enzyme, and Chymase Gene Polymorphisms as Biomarkers for Basal Cell Carcinoma Susceptibility.

The intake of angiotensin-converting enzyme (ACE) inhibitors and specific antagonists of angiotensin II receptors, widely used as antihypertensive drugs, significantly reduces the risk of developing basal cell carcinoma (BCC), highlighting the possible tumorigenic role of angiotensin II (AngII). We present here the investigated genetic association between the development of BCC and functional DNA polymorphisms M235T, I/D, and A1903G in the genes of angiotensinogen (AGT), angiotensin-converting enzyme (ACE), and chymase (CMA1), which mediate AngII production levels. DNA samples of 203 unrelated Greeks were studied, including 100 patients with BCC and 103 matched healthy controls. The MT genotype of the AGT-M235T polymorphism was significantly more prevalent in the patient group (78.0%) versus the healthy control group (28.3%; p<0.001). The DD genotype of the ACE-I/D polymorphism was also increased in BCC patients (72.8%) compared to controls (46.2%; p=0.001). The heterozygous AG genotype of CMA1-A1903G was significantly more frequent in the BCC group (86%) than in the healthy controls (50.5%; p<0.001). The MT, DD, and AG genotypes of the AGT- M235T, ACE-I/D, and CMA1-A1903G polymorphisms, respectively, were significantly increased in frequency within the group of cancer patients compared to the healthy controls. All three genotypes correspond to increased enzyme levels or activity and result in increased levels of AngII; therefore, they may be potentially utilized as reliable biomarkers associated with an individual's increased risk for BCC development.

Оценка состояния генофондов популяций Pinus sylvestris L. на востоке и северо-востоке Восточно-Европейской равнины

A molecular genetic analysis of horologically related populations of Pinus sylvestris L. was carried out. In the east and northeast of the East European Plain using intermicrosatellite analysis of DNA polymor-phism. Data on the genetic diversity of populations were obtained and the state of their gene pools was assessed. The studied populations of Scots pine have a high genetic diversity (P95= 0.938, HЕ= 0.170, ne= 1.540). Of the 144 DNA fragments, only 3 (0.021%) are rare. The analysis of the share of rare alleles showed that the genetic structure is less balanced in the populations of Verkhnevetluzhskaya (h= 0.254) and Vetluzhsko-Vyatka (h= 0.273). The most balanced genetic structure was observed in the Moloma population (h=0.112) and in the North Vyatka-Uvala population (h=0.127). When assessing the state of gene pools, it was found that the highest values of the coefficient of genetic originality (CSR) were found in the populations of Sysolo-Vychegodskaya (1,164) and Volga-Vetluzhskaya (1.140). This indicates the high specificity of the gene pools of these populations. The lowest values of CLO were determined in the Vetluzhsko-Vyatskaya (0.857) and Volzhsko-Surinskaya (0.875) populations. Analysis of all three groups of indicators of the state of gene pools of populations showed that, taking into account the genetic structure and genetic originality, 6 populations of P. sylvestris have a satisfactory state of gene pools, and two populations (Verkhnevetluzhskaya and Vetluzhsko-Vyatka) have depleted gene pools. When selecting trees for conservation and reforestation, it is necessary to preserve populations with both typical and specific gene pools.

Genetic diversity analysis and core collection construction for Amaranthus tricolor germplasm based on genome-wide single-nucleotide polymorphisms

• Genetic diversity among Amaranthus tricolor accessions was evaluated using genome-wide SNPs. • A set of 5638 SNPs without missing data in accessions was identified. • The population structure was analyzed among accessions. • A core collection of 105 accessions was created. The genus Amaranthus includes 60 to 70 species, some used as grain or leafy vegetable crops, others as ornamental crops. A. tricolor is used as a traditional leafy vegetable in Asia. It has a remarkably valuable nutrient composition and a high tolerance to environmental stresses but has been neglected in modern breeding. Detecting DNA polymorphisms among accessions helps in understanding the genetic diversity and population structure of germplasm collections. The genetic diversity and population structure were analysed among 465 A. tricolor accessions held by the World Vegetable Center genebank with genome-wide single-nucleotide polymorphisms (SNPs) developed by the double-digest restriction-site-associated DNA sequencing method. We found a total of 10 509 SNPs among the accessions and identified a set of 5638 SNPs without missing data in 440 accessions. This marker set was used to verify the species attribution of the germplasm, to analyse genetic diversity and population structure of the collection, and to create a core collection of 105 accessions representing the genetic diversity of 377 source accessions. This core collection is available for research and breeding (World Vegetable Center: info@worldveg.org). The SNP data generated here will contribute to exploring genetic loci related to economically and agronomically important traits in this species.

Biochemical and breakpoint cluster region-c-ABL oncogene 1 polymorphism study among Iraqi patients with chronic myeloid leukemia

BACKGROUND: Chronic myeloid leukemia (CML) has been well recognized as an exemplary instance of a malignant disease characterized by a distinctive molecular occurrence, namely the presence of the breakpoint cluster region (BCR)-c-ABL oncogene 1 (ABL1) oncogene. The Philadelphia chromosome gives rise to an anomalous fusion gene characterized by atypical kinase activity, resulting in the accumulation of reactive oxygen species and genetic instability that holds significance in the advancement of diseases. OBJECTIVE: The objective of this study was to investigate the detection rate of BCR-ABL1 polymorphism and BCR protein level in a group of Iraqi patients with CML. MATERIALS AND METHODS: This study has been carried out on 150 specimens, 120 patients subjected to CML included 20 patients diagnosed as newly diagnosis CML and 100 patients treated with CML. In addition to 30 apparently healthy persons as a control group (normal persons) from the National Center of Hematology/Mustansiryiah University/Baghdad, 65 out of 100 patients on imatinib while 35 nonimatinib (nilotinib and bosutinib). Fresh whole blood and serum were obtained from all patients and controls. We used total DNA genomic extraction extracted from ethylenediaminetetraacetic acid blood for genetic detection of Bcr/Abl Genes Polymorphism by sequencing technique in patients with CML and apparently control groups and used serum for biochemical tests include urea, lactate dehydrogenase (LDH), aspartate transaminase (AST), alanine transaminase (ALT), and creatinine using biochemicals methods (colorimetric and kinetic), respectively, as well as detection BCR protein level using sandwich enzyme-linked immunosorbent assays technique. RESULTS: According to age and sex, the patients’ groups were matching with the control group. Regarding the biochemical parameters (urea creatinine, ALT, AST, and LDH) serum level, there are no significant differences among new diagnosis CML, patients respond to treatments and failure group except in serum level of creatinine between new diagnosis CML group and failure group, there are significant differences (P = 0.01). The present results showed that DNA polymorphism distribution was according to C\C; G\C; A\T; and A\A were 32%, 26%, 18%, and 24%, respectively, in patients with CML and 28%; 20%;12%; and 40%, respectively, in the control group. There are significant statistical differences (P &lt; 0.05) between different groups according to the genotyping of BCR\ABL, the results obtained from the sequenced 429 bp fragments, and the detailed positions of the observed variations are described in the NCBI reference sequences (rs766724113). The samples were submitted in NCBI, and the accession number of nucleotide sequences of BCR\ABL as new recording: LC 775148, LC 775149, and LC 775150, while regarding with BCR protein, there are significant differences in level between new diagnosis CML and CML on treatment and control groups, P &lt; 0.001 for each comparison while there are no significant differences between treated group and control group (P = 0.729). CONCLUSION: The present results indicate that BCR-ABL1 polymorphism and BCR protein level in a group of Iraqi patients with CML may play a role in the tumor biology of the examined subset of CML and may contributed to their development.

Analysis and evaluation of correlation between DNA polymorphism in the genes MTHFR, PAI-1 and serum creatinine, creatinine clearance and albumin/creatinine ratio in morning urine of patients with type 2 diabetes mellitus and diabetic nephropathy.

Diabetic nephropathy is a major microangiopathic complication of type 2 diabetes and a leading cause of chronic kidney disease (CKD).

Molecular Characterization of the Honeybee (Apis mellifera) from two Vegetational Zones in Nigeria

The genetic structure and diversity of Apis mellifera from eight populations across two vegetational zones of Nigeria was investigated in order to provide base line information that will enhance its conservation, genetic improvement and sustainable yield. Genomic DNA was extracted from 40 specimens randomly selected from eight colonies in derived savannah and tropical rainforest regions of Nigeria. The specimens were amplified using five Random Polymorphic DNA (RAPD) primers and the amplicons generated were assessed using appropriate statistical indices. Results generated revealed significant genetic polymorphisms among the eight populations of A. mellifera ranging from 38.89 to 68.52% with a total of 281 bands amplified from 54 loci in all samples. The derived savannah population (Offa) had a greater amount of genetic diversity than the rest seven (7) populations as revealed by the percentage of polymorphic loci, expected heterozygosity and Shannon information index (I). The genetic structure as revealed by the Neighbour-Joining dendrogram showed that the eight populations studied represent two major genotypic groups with intra-group relationships. Eleyoka and Offa populations were the genetically closest in the first group while Ayetoro-Gbede and Ayegunle-Gbede populations were the genetically closest in the second group. The study suggests that the two major genotypic groups represent two distinct genetic stocks and can thus be managed accordingly.

Diversity Analysis of Rhizoctonia solani Kuhn Isolates Causing Sheath Blight of Rice in Eastern Uttar Pradesh, India

The sheath blight of rice is a devastating fungal disease, which is caused by Rhizoctonia solani AG1 IA. Twenty-one sheath blight isolates of rice were collected from different locations in eastern Uttar Pradesh, India, to study the variation in cultural, pathogenic, and molecular characterization. All the R. solani isolates were classified into four groups depending on the arrangement of sclerotia, i.e., peripheral, centre, scattered, and last group includes placing at the centre and peripheral 11, 1, 5, and 4 isolates, respectively. Depending on pathogenicity, isolates are grouped into weakly virulent, virulent, and highly virulent, representing 12, 4, and 5 isolates, respectively. The random amplified polymorphic DNA has been successfully used for molecular characterization. In our study, ITS1/ITS4 and AG-1 IA-specific markers yielded reproducible banding patterns. UPGMA cluster analysis revealed two major clusters, A and B, with a 13.85% dissimilarity value. This study does not correlate with virulence, geographical location, and RAPD profile groupings.

MOLECULAR, PHENOTYPIC MARKER ASSAYS, AND RADIOSENSITIVITY TESTS OF GAMMA-IRRADIATED CELOSIA ARGENTEA

Random amplified polymorphic DNA (RAPD), inter simple sequence repeat (ISSR), and UPOV phenotype markers were used to study the DNA polymorphism in gamma-ray induced morphological mutants of Celosia argentea var. plumosa Hungarian variety ‘Arrabona.‘ In the experiments, the study determined the radio sensitivity and the genetic diversity of gamma radiation of C. argentea var. plumosa ‘Arrabona.‘ Seeds of C. argentea var. plumosa ‘Arrabona‘ were irradiated with gamma rays to increase their genetic diversity. The irradiation doses consisted of 0, 75, 150, 300, 450, and 600 Grays (Gy). The germination percentage, survival rate, and phenotype of irradiated plantlets underwent evaluation in the first (M1) and second (M2) generations. The investigation of genetic diversity used the ISSR and RAPD primers. Based on the results, the first-generation genetic distance increased as the doses increased. But the trend changed considerably through the generation due to the low condition and fertility of the high doses of gamma-irradiated plants. These individuals did not show at the next mutant generation, changing the population gene pool. In addition, open pollination has also changed genetic diversity. The RAPD and ISSR primers proved proper to evaluate the genetic diversity, nonetheless fewer direct connection occurred between the appearance and the used RAPD or ISSR markers. The LD50 dose between 150 and 300 Gray treatments and the radiation between 300450 Gray induced the median growth reduction in the mutant ‘Arrabona‘ population. Based on these results, the study concluded that both UPOV-based phenotyping and molecular marker analysis revealed appropriate for determining genetic divergence, but detecting greater genetic distance resulted in molecular markers.

Molecular Characterization of Different Soybean [Glycine max (L). Merril] genotypes by RAPD Markers

The present investigation was carried out on molecular characterization of different soybean genotypes by using RAPD markers. Genetic diversity of 10 soybean genotypes was studied using Random Amplified polymorphic DNA (RAPD). 12 RAPD primers were used for screening 10 soybean genotypes from which total 334 fragments were amplified. It was observed that 41.10% bands were polymorphic and 58.88% were monomorphic. The percent of polymorphic amplicons in banding pattern was calculated and it was highest in OPA-01 (67.85%) followed by OPA-11 (66.66%). Polymorphism information content (PIC) was calculated and highest PIC value was observed in OPA-01 (0.26) and lowest was observed in OPA-08 and OPA-05 (0.03). Maximum amplicons were produced in OPA-01 (56) and minimum in OPA-08 (11). Genetic relationship between soybean genotypes was determined on the basis of Jaccard’s pair wise similarity coefficient values and dendrogram was generated by UPGMA (Unweighted Pair Group Method with Arithmetic Mean) cluster analysis using dice’s similarity matrix through NT-SYS pc software. The dendrogram of RAPD analysis shows 8 clusters and sub clusters were revealed. The value of similarity coefficient ranged from 0.19 to 0.71. The maximum similarity percentage i.e. 71% was found between KDS-726 and KDS-922 genotypes. The minimum similarity i.e. 19% was found between IVT-33 and IVT-13. According to this information it is concluded that the KDS-726 and KDS-922 genotypes showed minimum genetic diversity as compare to the IVT-33 and IVT-13 genotypes. Therefore, understanding the genetic diversity of soybean genotypes is essential to broaden the genetic base and to further utilize in the MAS breeding program.

RAPD Typing, Antibiotic Resistance Profiling and Genetic Diversity of Candida Isolates

Candida species is a leading cause of systemic mycosis in hospitalized patients, as well as morbidity and mortality globally. The purpose of this study was to observe the genetic diversity of Candida species among different soils located in different regions of Mumbai Suburban and their drug resistance pattern. A total of 86 soil samples were screened by using Candida Differential Agar and 19 isolates were obtained. Molecular analysis of these isolates revealed that 14 Candida albicans were the most common species, followed by 3 Candida tropicalis and 2 Candida krusei. The drug resistance pattern was studied by performing an Antifungal Susceptibility Test (AST) and determining Minimum Inhibitory concentration (MIC). The isolates did not show much variation in their resistance profile and were all resistant to antifungal drugs except, Amphotericin B. Candida tropicalis was highly resistant to Amphotericin B withstanding a concentration of 500μg/ml. Random amplified polymorphic DNA of Candida species showed that these species were highly similar to each other except for Candida tropicalis which showed a divergence while the other species were in the pair according to dendrogram analysis. The findings of the current study revealed genetic diversity and a correlation between antifungal susceptibility profiles and the genotype groups of Candida species isolated from different soil samples. We also found that Amphotericin B and Voriconazole were the most effective antifungal drugs against different Candida species.

RAPD MARKER TO SCREENING GENETIC DIVERSITY OF LOCAL CHICKEN

A total of 500 local chickens from different regions of Iraq, which were divided into five groups according to the regions from which they were taken, namely, the East (E), West (W), North (N), South (S), and Central (C) regions of Iraq. The blood samples were collected into tubes containing an EDTA, DNA extraction was carried. Using 20 primers from Gen Script USA, random amplification of polymorphic DNA (RAPD-PCR) was carried out. The reaction of PCR had a final volume of 25 μl. A total of 473 loci ranging between 200 and 1800 bp were amplified. The higher numbers of bands shown in north(N) local chicken group (101) than the other groups. East (E) and South (S) groups phenotypic revealed higher numbers of the polymorphic bands. The Polymorphism percent (%6.2) in East (E). Between the north (N) and west (W) groups, the genetic similarity score was found to be the highest (0.893). dendogram It was clear that the South(S) appeared to be most distant from the other groups whereas the West (W), North (N) were related closely together whose genetic closeness is highest. The main aim of present study is to assess the genetic specificity chickens in Iraq independs on RAPD analysis, To assess phylogenetic relatedness between groups of chickens, is to understand the extent of genetic variation between local chickens groups in Iraq.

In Vitro Activity, Stability and Molecular Characterization of Eight Potent Bacteriophages Infecting Carbapenem-Resistant Klebsiella pneumoniae.

Members of the genus Klebsiella are among the leading microbial pathogens associated with nosocomial infection. The increased incidence of antimicrobial resistance in these species has propelled the need for alternate/combination therapeutic regimens to aid clinical treatment, including bacteriophage therapy. Bacteriophages are considered very safe and effective in treating bacterial infections. In this study, we characterize eight lytic bacteriophages that were previously isolated by our team against carbapenem-resistant Klebsiella pneumoniae. The one-step-growth curves, stability and lytic ability of eight bacteriophages were characterized. Restriction fragment length polymorphism (RFLP), random amplification of polymorphic DNA (RAPD) typing analysis and protein profiling were used to characterize the microbes at the molecular level. Phylogenetic trees of four important proteins were constructed for the two selected bacteriophages. All eight bacteriophages showed high efficiency for reducing bacterial concentration with high stability under different physical and chemical conditions. We found four major protein bands out of at least ten 15-190 KDa bands that were clearly separated by SDS-PAGE, which were assumed to be the major head and tail proteins. The genomes were found to be dsDNA, with sizes of approximately 36-87 Kb. All bacteriophages reduced the optical density of the planktonic K. pneumoniae abruptly, indicating great potential to reduce K. pneumoniae infection. In this study, we have found that tail fiber protein can further distinguished closely related bacteriophages. The characterised bacteriophages showed promising potential as candidates against carbapenem-resistant Klebsiella pneumoniae via bacteriophage therapy.