فصلنامه دنیای میکروب ها
سال یازدهم شماره 4 (پیاپی 37، زمستان 1397)

Tuberculosis has become one of the most serious diseases due to theincreased prevalence of resistance. To control the disease, tuberculosis molecular epidemiological studies are important in determining the genetic diversity of strains circulating in communities. The aim of this study was to investigate the molecular epidemiology of drug-resistant tuberculosis among patients in Iran and Azerbaijan.   The isolates from tuberculosis patients of the Azerbaijan Republic andTabriz were studied from March 2014 to March 2015. All isolates were identified as Mycobacterium tuberculosis complex by biochemical methods and drug susceptibility testing against isoniazid, rifampicin, streptomycin, and ethambutol was performed by proportional method. Following DNA extraction, each MIRU locus was amplified individually to determine their repeat numbers as compared to H37Rv standard strain. Finally, genotyping was performed according to the repeat numbers of subsets of 15 studied loci using the MIRU-VNTRplus reference website.   Totally of 119 obtained isolates, resistance to at least one drug of the first line of anti-TB drugs and multidrug resistance were observed as 27.73 and 6.72 percent, respectively. Cameron, UgandaӀ, UgandaӀӀ, LAM, TUR, Dehli/CAS, Bovis, Beijing, NEW-1 and H37Rv were among genotypic families which were determined.   According to the results of this study, the rate and pattern of drug resistance andgenotypic families are different among the people of Tabriz and Azerbaijan, but the indicators were more diverse in Azerbaijan as compared to Tabriz.

Hepatitis C virus is the main cause of chronic hepatitis, resulting in a thousand deaths every year. NS5B protein is an RNA-dependent RNA polymerase that is encoded by the NS5B gene and involved in virus replication. One of the most effective drugs in the treatment of infections caused by this virus are NS5B protein inhibitors. The emergence of the strains resistant to these drugs is a major obstacle to the success of treatment. The aim of this study was to investigate possible mutations in the NS5B region of hepatitis C virus genotype 1a in Guilan province.   Materials & HCV genomic RNA was extracted from the plasma samples of 225 anti-HCV positive patients and its molecular identification and genotyping was carried out by RT-PCR and product Sequencing. Then, the NS5B gene was amplified in 10 strains with genotype 1a and subsequently sequenced to determine mutations in this region.   Genotypes 3a (53.3%) and 1a (36.9%) were the most abundant genotypes identified in Guilan. According to sequencing results, out of 10 investigated genotype 1a strains, 5 strains showed 7 types of missense mutations in codons Q309, A327, S254, K304, N307, R250, and A334.   Genotype 1a is one of the common genotypes of hepatitis C virus in Guilan. Identifying mutations or polymorphisms associated with resistance in hepatitis C virus can be useful in optimizing the treatment and determining the efficacy of drugs in treating hepatitis C virus.

Bacterial cellulose synthesized by some microorganisms, including Acetobacter xylinum, has been widely used in various industries due to its specific properties. The purpose of this study was to optimize the cultivation condition for the production of microbial cellulose in a new culture medium.   Materials & In this experimental study, new sources of carbon and nitrogen were added to the Hestrin-Schramm medium, containing A. xylinum, and incubated for 7 days under static conditions. Carbon sources included glucose, galactose, fructose, lactose, sucrose, maltose, ethanol, methanol, inositol, glycerol, xylose, and mannitol and nitrogen sources included ammonium sulfate, ammonium nitrate, sodium nitrate (1, 3, 6, 9  g/l HS medium), peptone and yeast extract (5, 10, 15, 20  g/l  HS medium). Sodium alginate and sodium acetate were used to investigate the viscosity effect and to adjust the medium pH. Scanning electron microscopy, X-ray diffraction, and FTIR spectroscopy technique were used in order to confirm the cellulose production. Sodium alginate and sodium acetate were used to investigate the viscosity effect and determine the pH adjustment. Scanning electron microscope, X-ray diffraction and FTIR spectroscopy technique were used in order to confirm the cellulose production.   Four carbon sources including glycerol (without a significant drop in pH), glucose, fructose, and inositol produced the highest amount of cellulose, respectively. Organic nitrogen sources, particularly peptone, had a great impact on cellulose production, unlike mineral nitrogen sources. The optimum amount of sodium alginate as the viscosity agent and sodium acetate as the buffer was 1.2 and 3 gram per liter of culture medium, respectively. X-ray diffraction showed the highest crystallinity index in medium containing glucose, fructose, inositol, and glycerol, respectively. The amount and intensity of infrared absorption in FTIR scanning of the products of culture media containing glucose and glycerol and comparing them with other similar cellulose graphs confirmed the cellulose production. Furthermore, Scanning Electron Microscopy studies clearly showed a nanofiber structure of microbial cellulose in media with better carbon sources.   According to our findings, glycerol and peptone have the most impact on microbial cellulose production. It was also indicated that addition of 2.1 g/ L sodium alginate to the culture medium as the viscosity agent along with pH control during the process by adding 3 g/ L sodium acetate can have a significant effect on cellulose production.

Poly-aromatic hydrocarbons are one of the most important toxic compounds of crude oil and environmental pollutants. The purpose of this study was to isolate halo-tolerant pyrene- degrading yeast.   Isolation of yeasts from contaminated and saline soils of oil-rich southern regions was carried out, at first. Then, screening of yeast isolates was performed based on their ability to grow in the salty medium and biodegrade oil. After screening and molecular identification of the optimal strain, its ability to tolerate salt and grow in the presence of pyrene, as well as the ability to degrade pyrene and other low molecular weight hydrocarbons were studied.   In this study, EBL-C16 was selected as the superior isolate, with growth in salt concentrations of 0-15% and eliminating 75.51% of the crude oil. Molecular identification of this isolate showed 100% similarity to Basidioascus persicus. Growth analysis showed the ability of this yeast isolate to grow in salt concentrations of 0 to 20 %. Growth and removal studies in the presence of 500 mg/l of pyrene and 2.5% salt showed 78.57% pyrene removal after 21 days, with the growth rate of 1.4 g/l of dry weight, as well as CO2 production of 3.1 mg. B. persicus had the ability to break down phenanthrene and anthracene, as well.   The results can be used to use halo-tolerant yeasts for bioremediation of oil-contaminated saline areas.

Sulfide compounds of ceramic industries wastewater cause water pollution as well as plants and aquatic destruction.  This study was aimed to evaluate sulfide compounds removal from ceramic industries wastewater by Thiobacillus thioparus and indigenous wastewater bacterial isolates.   Materials & Indigenous bacterial strains were proliferated at pH of  7, the temperature of 25oC, agitation speed of 200 rpm and an aeration rate of 100 mL min-1 in a 2 L bioreactor for 15 consecutive cycles. Sulfide compounds removal function of T. thioparus and indigenous bacterial strains along with the effect of pH and initial sulfide concentrations were investigated.   The results showed a thiosulfate removal rate of 250 mg sulfide L-1 h-1, a thiosulfate conversion percentage of 100% and a thiosulfate oxidation time of 44 min following 8 consecutive cycles. The sulfide removal rate of T. thioparus and ceramic wastewater indigenous bacteria was obtained as 246.5 and 276.5 mg sulfide L-1 h-1, respectively. Sulfide removal rate by proliferated bacteria decreased from 250 at pH of 7 to 230 and 180 mg sulfide L-1 h-1 at pH of 8 and 9, respectively. Bacterial isolates had an acceptable function in sulfide concentration of 3000 mg L-1, as well. Sulfide removal ability of T. thioparus isolates was decreased by 2.5 and 4 folds, when pH changed from 7 to 8 and 9, respectively. This bacterial strain was not able to tolerate high sulfide concentrations.   The results showed that bacteria isolated from ceramic industries wastewater have a higher capability of sulfide compounds removal as compared to T. thioparus isolates.

According to the FAO annual report, 10 percent of the world's food products are contaminated with fungal toxins, among which aflatoxins have the most contribution as compared to others. This research was aimed to assess the ability of Lactobacillus rhamnosus strain GG in the reduction of aflatoxin B1 (AFB1) in a simulated human gastrointestinal tract containing sterilized milk.   For this purpose, the bacterial count and aflatoxin concentration were adjusted to 1×1010 cfu/ml and 5 ppm, respectively, where artificial gastrointestinal tract discharges were inoculated in the simulated environment. In this study 6 treatment groups were assessed in the presence and absence of bacteria, sterilized milk, and gastrointestinal juice suspension. The concentration of residual aflatoxin was determined using high-performance liquid chromatography (HPLC) and purification by Immunoaffinity column.   The reduction of aflatoxin B1 at all treatments was determined using HPLC with a detection limit of 0.25 mg/ml and a quantification limit of 0.75 mg/ml. The mycotoxin recovery rate was between 89% and 94% for AFB1. The aflatoxin B1 calibration curve with a correlation coefficient of 0.95 was linear in the concentration range of 1 to 10 ng/ml. The highest and lowest average removal percentage of aflatoxin B1 was observed for 5 and 1 treatments (58.8 ± 0.018 and 13.86 ± 0.017) respectively, where a significant difference in removal percentage was observed among six treatment groups.   The results indicated that beside L. rhamnosus strain GG, gastric and small intestine juices are suitable to eliminate or reduce aflatoxin B1, as well.

Eggplant fusarium wiltis an important factor of yield reduction throughout the world. The ability of this pathogen to survive for several consecutive years within the soil, even in the absence of the host, has made it difficult to control. Producing and using the resistant cultivar is the most effective and suitable method to control this disease. This study was aimed to identify fusarium wilt resistant eggplant cultivars. First, leaf samples of domestic and hybrid eggplant cultivars were gathered from 28 provinces in Iran and then DNA extraction from young leaves of the cultivars was carried out using CTAB method. Four markers including CAPS, RAPD, SRAP, and SCAR were used to determine the resistant cultivars. In order to confirm the results, resistance and sensitivity of the genotypes were assessed in greenhouse conditions, as well.   Out of 20 genotypes of this study, 13  showed index resistance band using CAPS, RAPD, and SRAP of molecular markers. On the other hand, the SCAR marker could not separate the resistant cultivars from the sensitive ones. Phenotype assessment of native and hybrids resistant cultivars in greenhouse condition confirmed the results of the molecular analysis.   In general, the use of resistant cultivars obtained in this study using molecular markers is recommended for planting in areas with fusarium wilt disease.

Soft rot is one of the most important bacterial diseases of potato in Iran and the world. The aim of this study was to investigate the phenotypic and genotypic diversity of these agents in the main potato- growing regions in Iran. Samples were collected from tubers that were suspected of soft rot disease during harvest. Phenotypic characteristics were assessed by conventional methods. Genetic diversity was determined using genomic fingerprinting techniques. Cluster analysis and population structure analysis were performed by the SplitsTree 4.11.3, Arlequin 3.11 and the STRUCTURE 2.3.4 software.   The study showed a high level of diversity among isolates so that some of them showed phenotypic characteristics that differed from the isolates described in previous studies. Studied isolates were placed into two major groups of Pectobacterium wasabiae and Pectobacterium carotovorum subsp. carotovorum. Genomic fingerprinting profiles revealed that the isolates are genetically heterogeneous and classified into three genetic populations. 50.0% of the phenotypic group 1 isolate and 77.3% of the phenotypic group 2 isolates belonged to genetic population 1 and 3, respectively. No correlation was observed between genomic fingerprints, geographic areas, and potato cultivars,  but the genetic distances and the maximum and minimum gene flow were correlated with geographic distance.   According to the results of this study, seed trade may introduce soft rot disease to new territories in the neighboring provinces.