Iraqi Isolates Research Articles

Leishmanicidal activity of Artemisinin against cutaneous Leishmaniasis, in Vitro

Background: Cutaneous leishmaniasis (CL) is a neglected disease in tropical countries, including Iraq. Several studies have sought to examine chemotherapies for leishmaniasis treatment but most of them are of toxic and/or undesirable side effect, therefore, the need for investigating new fewer toxic therapies is essential.
 Aim of study: In this study, the cytotoxic effect of Artemisinin (ART), a novel herbal compound, was screened against the two forms, promastigotes and amastigotes, of the Iraqi isolate of Leishmania tropica, the causative agent of Baghdad boil.
 Material and methods: Different concentrations (1000, 500, 250, 125, 62.5, 31.25, 15.6 and 7.8) µM of Artemisinin were screened to investigate the leishmanicidal activity of the herbal compound against the two forms of the parasite along three times of follow up (24, 48, 72) hour using MTT cytotoxicity assay.
 Results: The results showed that growth rate and cell viability were significantly decreased at all studied concentrations. The IC50 was measured after 72 hours of follow up and was 2.625 µM and 2.636 µM for promastigotes and amastigotes, respectively.
 Conclusion: These findings approved the leishmanicidal efficacy of Artemisinin against the of L. tropica and can be further studied to screen its effectiveness in vivo for exploring a safer herbal drug for treatment of cutaneous leishmaniasis.

Molecular Detection of wzx1 and wzy Genes in Multi Drugs Resistance E. coli Isolates

Escherichia coli have been genetically changes in Iraqi environments, the present study was carried out to detection E coli isolates sero-group using o-antigens gene cluster, tow genes were amplified used multiplex PCR, the results of present study show that present isolates have more than one plasmid in different size (500-700 bp). PCR results show about 100% of isolates were had a positive results of PCR products of O45 wzx1 gene (451 bp) while 25% of isolates were a positive PCR products of O45 wzy1 gene (255 bp) and about 25% of isolates have tow genes. A new finding in present study that 50% of isolates have other copy of O45 wzx1 gene, these results concluded that it may be important in Iraqi isolates identification and classification however we need more information about genes sequencing.

MOLECULAR DETECTION AND ANALYSIS OF MUT L GENE FROM Pseudomonas aeruginosa IN URINARY TRACT INFECTION PATIENTS

This study was aimed to detect and analysis of mutL gene from Pseudomonas aeruginosa in UTI patients .Two hundred midstream urine specimens were collected from UTI patients. Laboratory diagnosis carried out depend on morphological and biochemical tests and confirmed by using VITEK- 2 Compact system. The results showed twenty isolates of Pseudomonas aeruginosa from two hundred urine specimens in percentage (10%). Genome extracted from all twenty isolates by specific kit , then amplification of mutL gene in PCR by specific primers. Sequencing process of product PCR of all twenty isolates was done. MutL gene sequencing of all twenty isolates was alignment in NCBI and the phylogenetic tree was drew by using Geneious 10 software among sequence of locally isolates. The results of analysis sequence of mutL gene from locally isolates in phylogenetic tree showed seven different groups. Analysis comparison sequencing of mutL gene between locally and global isolates showed eight different groups in phylogenetic tree included three groups (A,G,H) identical with global isolates and five groups (B,C,D,E,F) were not identical with global isolates , therefore selected one isolate (SD-9) from these five locally groups and documented as anew Iraqi isolate in national GenBank and accepted under accession number (LOCUS MF898429) of nucleotides sequence and protein ID(AUI39222.1).

Effect of probiotic Saccharomyces boulardii in weight losing and cytokines levels in mice before and after Infection by Salmonella typhimurium

This study was designed to evaluate some immunological and nutritional effects of three types of Sacchromyces boulardii yeasts, which was, Mutant Iraqi Isolate (Mutant Saccharomyces boulardii) MSb, commercial strain (Commercial Saccharomyces boulardii) CSb and Local Iraqi Isolate (Local Saccharomyces boulardii) LSb, in the weight and the level of both interleukin-10 (IL-10), interferon-gamma (IFN-γ) in the mice orally administered with yeasts inoculum for 30 days before and after infected with Salmonella typhimurium. The results shown significant losing in the weights and significant increase in the level of IL-10, but there are no significant difference in the levels of IFN-γ at )P>0.05) in mice administered with MSb, CSb and LSb yeasts before infected with S. typhimurium. The results also shown significant difference in the weights, significant increase in the level of IL-10 and significant decrease in the level of IFN-γ at )P>0.05) in mice administered with MSb, CSb and LSb yeasts after infected with S. typhimurium. Despite the effectiveness of three species of yeasts in these parameters before and after infection, but MSb was the most efficiency.

The Effect of Coumarin Derivatives(compounds) on the Vibrio cholerae Isolates from Different Clinical Iraqi Sources

From a large number of bacterial samples collected from different hospital in Iraq in central health laboratory ,only ten isolates were identified primary as Vibrio. A number of morphology and biochemical test were carried out to complete this identification that showed all bacterial isolates were related to Vibrio cholerae .In this study all Vibrio isolates were investigated for Bio typing and the result showed that all (10) isolate were related to (Eltor biotypes) .Also, the susceptibility test towards eight antibiotics were carried out . Results shows that ciprofloxacin , Norfloxacin, Erythromycin, Ampicillin, ceftriaxone and Amikacin were the most effective antibiotics and their resistance percentage were 20%,20%,20, 20,30% and 30% respectively ,While Chloramphenicol and Co- trimoxazole were less effective and their resistance percentage were 90% both of them. Three (3,5,6) isolates V. cholerae were selected depending on results of antibiotics sensitivity tests as showed multiple –antibiotics resistance(100%). Then tested to study the effect of coumarin derivatives compounds (1, 2, 3 ) which showed inhibitory effect on V. cholera (3,5,6) isolates and the compound ( 3 ) showed the highest antibacterial activity of (12,15,14 mm) of inhibition zone diameter against V. cholera (3,5,6) isolates respectively. Also, these Iraqi isolates (3,5,6) used to test the effect of acridine orange (0.1%) as acuring agent , the results showed that all (3) isolates V. cholerae were sensitive to (ciprofloxacin, ceftrixone and Norfloxacin), While the rest were resistance to remained five antibiotics. The results of Agarose –gel electrophoresis of both normal V. cholerae (3,5,6) and cured isolates showed the presence of chromosomal and plasmid DNA bands in the normal case ,While only chromosomal DNA bands occur with V. cholerae (isolate 8) treated with an acridine orange at concentration of (10-2 to 10-4).

The Effect of Coumarin Derivatives(compounds) on the Vibrio cholerae Isolates from Different Clinical Iraqi Sources

From a large number of bacterial samples collected from different hospital in Iraq in central health laboratory ,only ten isolates were identified primary as Vibrio. A number of morphology and biochemical test were carried out to complete this identification that showed all bacterial isolates were related to Vibrio cholerae .In this study all Vibrio isolates were investigated for Bio typing and the result showed that all (10) isolate were related to (Eltor biotypes) .Also, the susceptibility test towards eight antibiotics were carried out .
 Results shows that ciprofloxacin , Norfloxacin, Erythromycin, Ampicillin, ceftriaxone and Amikacin were the most effective antibiotics and their resistance percentage were 20%,20%,20, 20,30% and 30% respectively ,While Chloramphenicol and Co- trimoxazole were less effective and their resistance percentage were 90% both of them. Three (3,5,6) isolates V. cholerae were selected depending on results of antibiotics sensitivity tests as showed multiple –antibiotics resistance(100%). 
 Then tested to study the effect of coumarin derivatives compounds (1, 2, 3 ) which showed inhibitory effect on V. cholera (3,5,6) isolates and the compound ( 3 ) showed the highest antibacterial activity of (12,15,14 mm) of inhibition zone diameter against V. cholera (3,5,6) isolates respectively.
 Also, these Iraqi isolates (3,5,6) used to test the effect of acridine orange (0.1%) as acuring agent , the results showed that all (3) isolates V. cholerae were sensitive to (ciprofloxacin, ceftrixone and Norfloxacin), While the rest were resistance to remained five antibiotics.
 The results of Agarose –gel electrophoresis of both normal V. cholerae (3,5,6) and cured isolates showed the presence of chromosomal and plasmid DNA bands in the normal case ,While only chromosomal DNA bands occur with V. cholerae (isolate 8) treated with an acridine orange at concentration of (10-2 to 10-4).

Molecular identification of Fusarium spp. isolated from tomato plant in Iraq and China

This study was conducted to identify Fusarium spp. isolated from tomato plant in Iraq and China. A total of 12 isolates from Iraq (1-12) and four isolates from China (M1-M4) were used in this study. Based on Morphological characteristics (color, growth pattern, Macro and Micro-conidia shape) high differences between Fusarium isolates were found. Sensitivity test to the fungicide carbendazim revealed that one third of isolates(4/12) from Iraq had EC50 values over than 1000?g/ml indicated that many Iraqi isolates have developed resistance to carbendazim. Based on ITS sequences, Fusarium isolates were identified as follow, isolates 1, 3, 5, 6, 7, 10 were identified as Fusarium oxysporum, isolates 8, 11 as F. solani, isolates 12, M1 and M3 as F. moniliforme, isolates 2, M2 and M4 as F. proliferatum, F. chlamydosporum and F. kyushuense , respectively.

Improved vaccine strategies of infectious bronchitis disease to reduce shedding of virulent virus from infected birds

Infectious bronchitis (IB) threatens the economies of entire nations, by adversely affecting the backyard farmers, poultry producers and poultry industry throughout the world. Infectious bronchitis virus (IBV), a member of the Coronaviridae family, is probably one of the most avian pathogens prevalent in the poultry industry worldwide. In the field which presents difference structures, especially in the S1 spike protein. Complicated immune response against IBV due to few similarities between serotypes. High mutation rate of the virus, in addition to the management and environmental factors, compromise the efficacy of the available vaccines and difficult the control of disease. Prepared effective IB vaccines from any IBV strain stimulates the immunity widely against morbidity and mortality after challenge with virulent IBV strains. On the other hand, vaccinates with live IBV vaccines do not reduce infection or viral shedding after challenge. In order to compare the prepared IBV vaccines taken four different IBV genotypes to determine stimulating immunity and the amount of viral shedding after challenge, two hundred fifty one day broiler chicks divided into five groups and vaccinated with oil-adjuvant vaccines prepared of Iraqi isolates (II, I-II, I- III and I-IV). different inactivated IB viruses including strains QX, CH Baghdad M 2014, CH Baghdad F1 2013, CH Babylon F2 2013, and an allantoic fluid control. Using a hemagglutination inhibition test for serum analysis after challenge with virulent IBV (Variant2) to knowledge antibody content against each of the vaccine antigens, Monitored the vaccinated birds daily for of morbidity and mortality were recorded, and used PCR to determine the amount of viral copy of examined clinical samples in selected periods of viral shedding. All vaccines used in the study gave good protection against morbidity and mortality, except the control group. Homologous vaccines gave good protection and lowest viral shedding in the trachea, kidney and feces compared to the heterogeneous vaccines.Key word: infectious bronchitis disease, inactivated vaccines, hemagglutination inhibition, viral load, challenge.

Isolation and identification of the Newcastle disease virus from field outbreaks in broiler and layer flocks in Iraq

Newcastle disease is one of serious pathological problems and causes of vast economic losses during 2011-2016 in Iraq. The disease caused high mortalities in all types of poultry nevertheless of vaccination. In this study all samples were collected from infected flocks with clinical signs of the disease. Inoculation of chicken embryonated eggs was carried out for virus isolation, identification, Haemagglutination and Haemagglutination Inhibition assay. Using Reverse Transcriptase-Polymerase Chain Reaction to confirm the presence of the virus, Intra Cerebral Pathogenicity Index and Mean Death Time were used to confirm all the isolates that were velogenic. The important determinant of Newcastle disease virus pathogenicity is fusion protein that has been used for phylogenetic analysis. sequencing and compared genetically of Newcastle disease virus Iraqi isolate to publish sequences acquired from GenBank showed 99% sequence similarity to the Iran isolate IRI 1392k (KJ176996.1). It can concluded from these data that introduction new virus was occurred in Iraq.

Effect of Soil Textural Classes on the Biological Nitrogen Fixation by Bradyrhizobium Measured by 15N Dilution Analysis

The current study was conductedas a pot experiment to determine the effect of soil texture on biological nitrogen fixation (BNF) of six most efficient local isolates, specified, of Bradyrhizobium. Cowpea (Vignaunguiculata L.), as a legume host crop, was used as a host crop and 15N dilution analysis was used for accurate determination of the amount of N biologically fixed under experimental parameters specified. Soils used are clay loam, sandy clay loam and sandy loam. Biological Nitrogen Fixation (BNF), in different soil textural classes, was as in the following order: medium texture soil > heavy texture soil > light textured soil. Statistical analysis showed that there is a significant variation in BNF % among six Iraqi isolates in the three soil textural classes. There is a significant variation in the number of the nodules of the six Isolates in one soil texture. However, nodules number does not agree with the BNF% in the same soil for any isolates. Statistical analysis of the data showed that there were significant differences in plant dry weight among the soil textural classes all over local isolates used in this study. Data also showed that there were significant differences in dry weight under different isolates.

Phylogenetic analysis and full-length characterization of S1 gene of IS-1494 (Variant 2) like infectious bronchitis virus isolates, Iran, 2015

Infectious bronchitis virus (IBV), a major pathogen of commercial poultry flocks, circulates in the form of different genotypes. Three IB viruses were isolated from broiler chickens showing respiratory and renal lesions. The isolates were characterized by reverse transcriptase polymerase chain reaction and sequence analysis of the Full-length of the S1. Three isolates were belonged to Variant 2 like (IS/1494 like) IBV genotype. Phylogenetic analysis showed that Variant 2 like isolates formed two clusters and the Iranian and Iraqi isolates were included in the cluster II. Cluster I composed of Israeli, Egyptian and Turkish Variant 2 like IBV Isolates. Three hyper variable regions (HVR) of S1 were determined. The Most variation was seen in HVR2. The findings emphasize the importance of continuous monitoring of IBV, in addition to adjust diagnostic methods, molecular epidemiological studies, development and use of vaccines which are adapted to the changing disease scenario.

Effect of Soil Textural Classes on the Biological Nitrogen Fixation by Bradyrhizobium Measured by 15N Dilution Analysis

The current study was conductedas a pot experiment to determine the effect of soil texture on biological nitrogen fixation (BNF) of six most efficient local isolates, specified, of Bradyrhizobium. Cowpea (Vignaunguiculata L.), as a legume host crop, was used as a host crop and 15N dilution analysis was used for accurate determination of the amount of N biologically fixed under experimental parameters specified. Soils used are clay loam, sandy clay loam and sandy loam. Biological Nitrogen Fixation (BNF), in different soil textural classes, was as in the following order: medium texture soil > heavy texture soil > light textured soil. Statistical analysis showed that there is a significant variation in BNF % among six Iraqi isolates in the three soil textural classes. There is a significant variation in the number of the nodules of the six Isolates in one soil texture. However, nodules number does not agree with the BNF% in the same soil for any isolates. Statistical analysis of the data showed that there were significant differences in plant dry weight among the soil textural classes all over local isolates used in this study. Data also showed that there were significant differences in dry weight under different isolates.

Molecular diagnosis of Newcastle disease Iraqi Virulent strain virus HN gene by specific primers design

Newcastle disease is considered the most contagious poultry disease and may cause severe economic loss in the poultry industry, The first report in which NDV was consider as zoonotic disease and a human pathogen that was published by Burnet, in 1943. The aims of this study was to detect HN gene of two NDV Iraqi isolates (Najaf APMV1/ Chicken/ Iraq-najaf/ ICCMGR/2012and Baghdad ICCMGR) by designing specific priemrs for the Iraqi isolates. DNV isolates propagated in embryonated chicken eggs and showed to kill embryos in different times intervals ( more than 40 hours and less than 72 hours) with marked sever hemorrhage in infected embryos and it was similar in the two isolates, agglutination activity reflect titer (1024) for Najaf isolate and (512) for Baghdad isolate. The present study successfully detected part of (HN) gene by using specific primers designed for one-step RT-PCR undertaken to use them on molecular diagnosis of the hemagglutinin-neuraminidase (HN) gene in the Iraqi isolates and can be used for NDV virus detection in Iraq.

Genotyping Based on AFLR1 Gene Sequencing in Local Iraqi Isolates of Phylopathoginic-Aflatoxigenic Aspergillus Flavus

Genotyping Based on AFLR1 Gene Sequencing in Local Iraqi Isolates of Phylopathoginic-Aflatoxigenic Aspergillus Flavus

Pathology of Experimental Encephalitozoon cuniculi Infection in Immunocompetent and Immunosuppressed Mice in Iraq.

This study was performed to evaluate pathology of experimental Encephalitozoon cuniculi (Iraqi isolate) infection in normal and immunosuppressed mice. Pathological changes were not seen in negative control mice while secondary bacterial infections were noted in the lungs, kidneys, and heart of mice given dexamethasone. Typical E. cuniculi infection lesions were found in brain, livers, lungs, and kidneys of mice given 107 E. cuniculi spores/mouse orally. These lesions were in the form of nonsuppurative meningoencephalitis with vasculitis in brain, interstitial inflammation with infiltration of both lymphocytes and plasma cells in lung tissue, and nonsuppurative interstitial (focal and diffuse) nephritis, presence of vacuole containing mature and immature spores in enterocytes within the tips of villi, and lymphoiod hyperplasia of the white pulp and vasculitis of the intratrabecular vessels. Mice that were given 107 E. cuniculi spores/mouse orally showed lesions similar to those observed in the previous group (vasculitis and granulomas) but the lesions were more severe and widespread. In conclusion, this is the first report of experimental E. cuniculi infection induced by E. cuniculi isolated from a naturally infected rabbit in Iraq and that infection became more severe and widespread upon the administration of dexaethasone.

Identification and characterization of a novel Iraqi isolate of Fusarium pseudograminearum causing crown rot in wheat

Crown rot is one of the main important fungal diseases affecting wheat in many areas of the world, including Australia, USA, and Iran. Until now, there had been no report of this pathogen in Iraq. Plants displaying crown rot symptoms were observed in Shaat Alarab (Basra, Iraq); we investigated the causal agent of the disease. Samples were surface-sterilized in bleach (1% available chlorine) and cultured on quarter-strength potato dextrose agar plates. DNA was extracted from fungal mycelia, using a modified CTAB protocol. The ITS/5.8S regions were amplified using primer pair ITS1 and ITS4. PCR products purified using a gel extraction kit were sequenced. The sequence that was detected was used to BLAST against NCBI data. The most similar sequence was the ITS/5.8S rDNA region of Fusarium pseudograminearum (strain NRRL28062), showing 97.95% identity. This species normally causes crown rot, resulting in severe damage under dry spring conditions. A pathogenicity test employed to assess the disease-causing ability of the strain showed significant disease symptoms up to 57% infected spikelets. The results confirmed the presence of F. pseudograminearum as a causal agent of wheat crown rot in Iraq. The presence of this pathogen demands further investigations to develop resistant cultivars and/or mechanical control.

Photodynamic Therapy for Acinetobacter baumannii Burn Infections in Mice

Multidrug-resistant Acinetobacter baumannii infections represent a growing problem, especially in traumatic wounds and burns suffered by military personnel injured in Middle Eastern conflicts. Effective treatment with traditional antibiotics can be extremely difficult, and new antimicrobial approaches are being investigated. One of these alternatives to antimicrobials could be the combination of nontoxic photosensitizers (PSs) and visible light, known as photodynamic therapy (PDT). We report on the establishment of a new mouse model of full-thickness thermal burns infected with a bioluminescent derivative of a clinical Iraqi isolate of A. baumannii and its PDT treatment by topical application of a PS produced by the covalent conjugation of chlorin(e6) to polyethylenimine, followed by illumination of the burn surface with red light. Application of 10(8) A. baumannii cells to the surface of 10-s burns made on the dorsal surface of shaved female BALB/c mice led to chronic infections that lasted, on average, 22 days and that were characterized by a remarkably stable bacterial bioluminescence. PDT carried out on day 0 soon after application of the bacteria gave over 3 log units of loss of bacterial luminescence in a light exposure-dependent manner, while PDT carried out on day 1 and day 2 gave an approximately 1.7-log reduction. The application of PS dissolved in 10% or 20% dimethyl sulfoxide without light gave only a modest reduction in the bacterial luminescence from mouse burns. Some bacterial regrowth in the treated burn was observed but was generally modest. It was also found that PDT did not lead to the inhibition of wound healing. The data suggest that PDT may be an effective new treatment for multidrug-resistant localized A. baumannii infections.