Iranian Journal of Microbiology
Volume:12 Issue: 6, Dec 2020

No abstract

Diphtheria is a potentially fatal disease caused by toxigenic bacterial infection, particularly from Corynebacterium diphtheriae (C. diphtheriae). Isolation of C. diphtheriae is technically lacking in sensitivity, and Elek’s test to detect toxin production has several difficulties associated with its application. Duplex real-time PCR to throat swab of suspected diphtheria patients can detect both bacteria and toxin-encoding genes simultaneously, faster, with higher sensitivity and specificity.

A total of 89 consecutive throat swabs from suspected diphtheria patients were collected from Sulianti Saroso Infectious Disease Hospital, Jakarta, during 2018 to 2019. Two pairs of primers and probes, targeting the rpoB gene of C. diphtheriae and the A-subunit of the diphtheria toxin gene, were used in this study. Parameters including annealing temperature, concentration of primers and probes, inhibitors, cross-reaction and detection limit were all optimized. Elek’s toxigenicity test and clinical data were analyzed for comparison.

The optimum annealing temperature was 55°C. The concentrations of Cd primer, Tox primer, Cd probe and Tox probe were 0.4, 0.6, 0.5 and 0.625 µM, respectively. DNA elution and template volumes were 50 µL and 5 µL. The detection limit was 2 CFU/mL. No cross-reaction with other microorganisms was observed. Of the 89 samples, duplex real-time PCR gave better results than the standard test, with 19 (21.3%) and 10 (11.2%) patients diagnosed with diphtheria, respectively.

Duplex real-time PCR increases the rate of laboratory diagnosis of diphtheria, compared to the standard method to detect potentially toxigenic C. diphtheriae.

Recent studies have hypothesized that sterile disc infection with the anaerobic Propionibacterium acnes, recently renamed Cutibacterium acnes, occurs in people with intervertebral disc (IVD) herniation. This study aimed to examine the presence of P. acnes in patients who have Low back pain (LBP) with Modic changes observed in their Magnetic Resonance Imaging (MRI).

Thirty-seven patients who were candidates for surgery due to disc herniation and demonstrated Modic changes in MRI were included in the study. Before the surgery, the level of pain in patients was assessed using the visual analog score (VAS). All patients were asked to fill in the Oswestry Low Back Pain Disability Questionnaire. Intervertebral disc changes observed in MRI were recorded for all patients. Then, during surgery, sterile intervertebral disc samples were taken. P. acnes detection was performed using PCR in the laboratory. Data analysis with Chi-squared test, independent samples t-test, and Mann-Whitney U test in SPSS 18.0.

The mean age of 37 patients equaled 43.64 years and the mean duration of symptoms was 11.05 months. In molecular examination, of the 37 individuals, the genome of P. acnes was positive in 23 cases (62.2%) and negative in 14 (37.8%). The relationship between VAS, disability score, changes in MRI, and patients’ age with the positivity of the intervertebral disc sample was also assessed. Of these variables, only age was significantly correlated with the positive molecular finding, such that with an increase in age, the probability of positive findings was increased (p = 0.022).

Based on the results, lumbar disc infection with P. acnes may play a significant role in causing Modic changes and the progression of the disease in patients with LBP.

Intestinal pathotypes of Escherichia coli belong to the companion animals may poses potential risk to public health following zoonotic transmission. Therefore, this study was proposed to determine the virulence genes associated to diarrheagenic E. coli strains isolated from healthy pet dogs and their owners in the southeast of Iran, Kerman province.

Totally 168 E. coli isolates were collected from 49 healthy household dogs and their owners. Seventy isolates were obtained from non-pet owners as control group. Presence or absence of the virulence genes including eae, stx1, stx2, st1, lt1, ipaH, cnf1 and cnf2 were screened by conventional polymerase chain reaction (PCR) and dissemination pattern of the genes were studied among the various hosts.

PCR examinations showed that the most frequent virulence gene was ipaH (6.1%) in dogs followed by eae in dog owners (6.1%) and in controls (8.6%). The most frequent pathotypes in dogs, their owners and controls were EIEC (6.1%), EHEC (4.08%) and EPEC (8.5%), respectively. In one of studied houses, both of dog and its owner harbored E. coli strains with same virulence profile (stx1/eae) and pathotype (EHEC).

These results collectively indicate that healthy household dogs probably are the mild reservoir of potential virulent E. coli strains with possible active transmission to their contact owner. However, even non-pet owners seemed to be a notable source of intestinal pathotypes, especially EPEC, for their environment. Transmission of E. coli pathotypes may occurs by direct contact with the reservoirs or ingestion of contaminated food. These pathotypes are potentially virulent and creates public health hazards. Further studies are needed for better understanding of dissemination mechanisms of E. coli pathotypes among humans and their pets.

Arcobacter species are food-borne and zoonotic enteropathogens. Defined breakpoints for the investigation of antimicrobial resistance of Arcobacter are missing.

The study was performed to investigate the incidence and antimicrobial resistance of Arcobacter species in animals and poultry meat samples procured from slaughterhouses in Iran. To investigate the prevalence of antimicrobial resistance, samples were collected from cattle (n=100), sheep (n=100), goat (n=100), broiler chicken (n=100), turkey (n=100) and quail (n=100). Arcobacter isolates of meat samples were isolated, investigated by PCR method and antibiotic resistance was also investigated. The susceptibility was assessed by Kirby-Bauer disc diffusion.

The results showed that 52 samples (8.66%) were positive for Arcobacter spp. The most prevalence were observed in broiler chickens (26%, n=26 samples), quail (13%, n=13 samples), turkey (8%, n=8), cattle (3%, n=3), sheep (1%, n=1) and goat (1%, n=1). Arcobacter butzleri had highest prevalence among Arcobacter species. All the isolates showed sensitivity to gentamicin, streptomycin and tetracycline.

Poultry meat is a potential source of infection with Arcobacter that must be considered in slaughterhouses in Iran. Arcobacter species showed sensitivity for a broad spectrum of antibiotics that can be used during infection with Arcobacter species.

The aim of present study was to evaluate the prevalence of Listeria monocytogenes and Escherichia coli, characterization and antimicrobial resistance of their serotypes and genotyping profiles in fresh beef and poultry meats marketed in Zanjan, Iran.

A total of 90 (45 chicken and 45 beef) samples were collected from January to June 2018 focusing on retail meat stores of Zanjan city, Iran. Foodborne pathogen detection and antimicrobial resistance of isolates performed by PCR and disc diffusion methods, respectively. Simplex PCR method was used for screening hly and uidA genes in L. monocytogenes and E. coli isolates, respectively.

Findings revealed high contamination in beef and chicken meats with E. coli (68.89% and 88.89%, respectively) and L. monocytogenes (53.33% and 46.67%, respectively). The most likelihood of E. coli isolates belonged to E. coli 13479 serotype. All L. monocytogenes isolates from beef and chicken meat samples had high similarity with serotypes L. monocytogenes strain NCTC 10357 and strain MF 4545, respectively. Multi drug resistance (MDR) was seen in both L. monocytogenes and E. coli isolates.

This study shows an insight of the current status of beef and chicken meat contamination maketed in Zanjan, Iran with E. coli and L. monocytogenes isolates (high contamination rate), their genotypic profile, epidemiological relationship and antimicrobial resistance (AMR) that should be considered as a significant public health concern in Zanjan, Iran.

Waste water from abattoirs could harbour bacteria some of which are pathogenic. Therefore, this study aimed to assess the quality of wastewater from some abattoirs in Ilorin, Nigeria.

The counts of viable bacteria, total coliform, faecal coliform, enterococci, S. aureus, P. aeruginosa and Salmonella/Shigella spp. of the wastewater was determined using selective media. The sanitary condition appraisal, antibiotic susceptibility test and plasmid profile of the isolates were assessed using standard methods.

The highest count of viable bacteria and total coliform obtained were 9.0 × 107 and 3.0 × 107 CFU/ml respectively. Faecal coliform and enterococcal count had the same highest value of 3.0 × 105 CFU/ml. The highest count of pathogenic bacteria: Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella spp. were 2.5 × 108, 1.9 × 107 and 3.0 × 104 CFU/ml respectively. The abattoirs sanitary scores ranged from 28.6-57.1%. The isolates showed multiple antibiotic resistance (MAR) index ranging from 0.5-1.0. Plasmid curing with 0.1 mg/ml of acridine orange solution led to reduction in the MAR index of most of the Gram negative bacteria. Pseudomonas stutzeri was susceptible to all the antibiotics while Proteus vulgaris was resistant to all the antibiotics after curing. Most of the Gram negative bacteria isolated belong to the families Enterobacteriaceae and Pseudomonadaceae while the Gram positive bacteria belong to the families Staphylococcaceae, Enterococcaceae and Streptococcaceae.

It was concluded from this study that wastewaters from the abattoirs were contaminated by bacteria with high MAR index. Most of these bacteria borne their antibiotic resistant factors in their plasmid.

The microbial contamination of wastewater is associated with health risks. The aim of this study was to use the autochthonous Bdellovibrio potential to prey Gram-negative pathogenic bacteria as a bio-control agent to treat urban wastewater.

Thirty-six raw sewage samples were collected for isolation of Bdellovibrio. Double layer plaque assay was used for isolation and the isolates were identified by microscopic examination and molecular analysis. To evaluate the predatory potential for decrease number of Gram-negative pathogenic bacteria, plaque perdition assay, reduction in host cells viability by colony-forming unit (CFU) counting, reduction in optical density (OD) in co-cultures and assay of killing efficiency were carried out. Also, the raw wastewater was treated by Bdellovibrio then the reduction in CFU counting and reduction in OD was evaluated.

Four strains of Bdellovibrio were isolated and were registered in Gene Bank. Clear plaques were observed after 3-6 days of incubation for all prey cells. The CFU enumerations of all preys were decreased after 48 hrs in co-cultures and raw wastewater. Also, OD was decreased down to 0.2 nm after 48 hrs.

These autochthonous Bdellovibrio strains are proposed to use for bio-control of Gram-negative pathogenic bacteria in wastewater and reuse it for irrigation in arid regions.

Tualang honey (TH) is a Malaysian multifloral jungle honey. In recent years, there has been a marked increase in the number of studies published in medical databases regarding its potential health benefits. The study aimed to investigate the effect of TH against Pseudomonas aeruginosa and Streptococcus pyogenes.

The effect of TH on both bacteria was investigated using MIC, MBC, growth curve, time-kill curve, scanning electron microscopy (SEM) and RT-qPCR.

The MIC of TH against P. aeruginosa and S. pyogenes was 18.5% (w/v) and 13% (w/v) respectively and MBC was 25% (w/v) for both bacteria. Spectrophotometric readings of at least 90% inhibition yielded MIC90 values of TH, 18.5% (w/v) and 15% (w/v) for P. aeruginosa and S. pyogenes respectively. A time–kill curve demonstrated a bactericidal with a 4-log reduction estimated within 8 hours. Using SEM, loss of structural integrity and marked changes in cell shape were observed. RT-qPCR analysis showed that TH reduced the pattern of gene expression in both bacteria, with a trend toward reduced expression of the virulence genes of interest.

This study suggests that TH could potentially be used as an alternative therapeutic agent for microbial infection particularly against these two organisms.

Essential oils (EOs) with different biological activities, such as antibacterial properties, are a valuable resource for developing new drugs.

Ingredients of six medicinally important EOs, including Artemisia dracunculus, Anethum graveolens, Citrus limon, Citrus sinensis, Cinnamomum zeylanicum and Zingiber officinale, were identified using GC-MS analysis. Moreover, their five major compounds were also listed. Furthermore, the half-maximal inhibitory concentration (IC50) against four important human bacteria was also investigated using the 96-well plate microdilution.

C. sinensis EO with IC50 of 1.0 and 4.7 mg.mL-1 have the most effect on the growth of S. aureus and P. aeruginosa. Moreover, EOs of Cinnamomum zeylanicum (IC50: 1.0 mg. mL-1) and Artemisia dracunculus (IC50: 1.3 mg.mL-1) significantly showed better inhibitory effect on E. coli and K. pneumoniae.

These EOs could be used for developing inexpensive, potent, and green antibacterial agents.

Gamma-aminobutyric acid (GABA) is a non-protein four-carbon amino acid that has many physiological properties, including reducing blood pressure, accelerating protein synthesis in the brain, and treatment of insomnia and depression. This amino acid is produced by a number of lactic acid bacteria, fungi and yeasts. The objective of the present study was to identify probiotic bacteria with the maximum ability to generate GABA and optimize the bacterial culture conditions having the highest potential for GABA production.

The potential of GABA production by Lactobacillus delbrueckii ssp. bulgaricus, Lactobacillus rhamnosus, Lactobacillus casei, Streptococcus thermophilus, Lactobacillus brevis and Lactococcus lactis ssp. lactis in the culture medium of MRS broth was assessed by High Performance Liquid Chromatography (HPLC). In order to increase the rate of GABA produced by the bacteria having the highest potential for GABA production, the conditions of the culture medium including pH (3.5 to 6.5) "temperature (25 to 45°C), time (12 to 96 h) and glutamic acid (GA) concentration (25 to 650 mmol) were optimized by the Box-Behnken’s Response Surface Method (RSM).

Lactobacillus brevis had the highest potential of GABA production (5960.8 mg/l). The effect of time and GA concentration was significant on the amount of GABA production. The best conditions of culture medium to achieve the highest amount of GABA production by Lactobacillus brevis (19960 mg/l) were temperature 34.09°C, pH 4.65, GA concentration 650 mmol and time 96 h.

The results showed that by optimization of the culture medium conditions of probiotic bacteria we can produce more GABA in culture medium.

Coenzyme Q10 is an anti-aging agent whose demand is increasing progressively. There are various strategies used for increasing coenzyme Q10 production by microorganisms. In this study, for the first time, we investigated the effect of iron oxide and silver nanoparticles on coenzyme Q10 production by Gluconobacter japonicus FM10.

In the first step, a preliminary experiment was set and carried out to obtain the minimum inhibitory concentrations of the nanoparticles on the strain FM10. Then the sub-MIC concentrations were used to investigate their effect on coenzyme Q10 production in the stationary and exponential phases of the growth, separately.

The results showed that coenzyme Q10 production increased in the presence of the iron oxide and silver nanoparticles. The silver nanoparticles induced 1.9 times higher coenzyme Q10 production. The highest level of coenzyme Q10 was induced when the silver nanoparticles were added to the culture medium at the stationary phase.

This should be noticed that so far nanoparticles have been considered as antibacterial agents, rather than being considered to cause probable beneficial effects on the induction of useful products in the microbial world. In this regard, their potential for increasing coenzyme Q10 production has received no attention. However, our present results showed that the nanoparticles can be used to increase the production efficiency of coenzyme Q10 in Gluconobacter.

Increasing the amount of protease from microbial sources is in the focus of attention. Random mutagenesis by physical methods like ultraviolet (UV) radiation is a cost effective and convenient procedure for strain improvement. Therefore, in the present study attempts were made to investigate the effect of UV radiation on Lysobacter enzymogenes in order to increase its protease activity.

UV mutagenesis was induced in L. enzymogenes fresh culture at the distance of 20 cm from light source for different exposure times of 70, 90, 150 and 200 seconds. The mutated isolates were randomly cultured from the nutrient agar medium to casein agar plate, as a selective medium. The primary screening was performed by observing hydrolysis of casein in the plate and the secondary screening was carried out on skim milk agar on the basis of zone of hydrolysis using bacterial supernatants. Quantification of protease activity was done by Anson’s method using tyrosine as standard.

UV radiation resulted in obtaining 12 mutants out of 100 examined L. enzymogenes strains with increased protease activity. The mutant M2, at 90s exposure time was selected as the best mutant bacterium which produced 1.96 fold more protease over the parent strain.

Random mutation by UV radiation is a simple and convenient method to increase the protease activity of Lysobacter enzymogenes. Furthermore, it seems that the middle time of exposure to UV, 90 s, was the best time because it can induce mutagenesis but did not hamper the bacteria growth and viability.

The great potential of bacteriophage for removing pathogen bacteria via targeting the cell wall is highly concerned. With a priority for drug-resistant, we screened endolysins  targeting Gram-negative bacteria to introduce a new antibacterial agent. The study was aimed to identify endolysins from the lysogenic phage of the Siphoviridea family in Bacillus subtilis.

The Bacillus subtilis strain DDBCC46 was isolated from a preliminary antibacterial screening program. The endolysin(s) was extracted, concentrated with ammonium sulfate saturation, and their activity evaluated against the indicator bacteria. The phage particles was extracted from the bacteria using the minimum inhibition concentration of mitomycin C, followed by testing the phage inhibitory effect on the growth of indicator bacteria. The NCBI, Virus-Host DB, and EXPASY databases were used to obtain and confirm the sequences of the genes encoding PG hydrolases in Siphoviridea phages hosted in B. subtilis.

An  816 bp gene encoding an endolysin enzyme, was approved in the B. subtilis, DDBCC 46, with specific primers of Bacillus phage SPP1. The purified-endolysin indicated antibacterial activity against Klebsiella pneumoniae, Salmonella typhimurium, Proteus (sp), and Escherichia coli. SDS-PAGE profiling followed by silica gel purification, led to introduce lys4630 as a therapeutic product and food preservative.

lys4630 showed specific effects on the very known gram-negative pathogens in clinics and food industries; E. coli, P. aeruginosa, Salmonella (sp).

Foodborne pathogens are among the serious problems all around the world and thus a novel and natural strategy to control and to inhibit such pathogens is highly demanded nowadays. The aim of this study was to isolate a specific bacteriophage of Escherichia coli O157:H7 from sewage in Fars province, Iran to determine its morphological and antimicrobial activities.

In order to isolate the bacteriophage of E. coli O157:H7, 10 samples of slaughterhouse wastewaters were used. Double-Layer Agar method was employed to isolate the bacteriophage. To identify the fine structure of the bacteriophage, electron microscope was employed. Host range and antibacterial activity of the phage was also investigated, in vitro.

The morphological and biological characteristics of a virulent Siphoviridae phage, PI, are reported. It was found that infection of E. coli O157:H7 strains with this specific bacteriophage produce clear plaques. In the one-step growth analysis, it was confirmed that the phage has been characterized with a very short rise period (around 15 min), an average burst size of 193 PFU/cell, high infectivity and potent lytic action. The bacteriolytic activity of PI was also investigated, in vitro. It was also clarified that at the MOI of 100, 10 and 1, the phage rapidly lysed the bacterial cells within 0.5 or 2 h.

These results indicate that the phage PI is a newly discovered phage against E. coli O157:H7 in Iran which may be recommended to use as bio-control purposes.

In a Turkish cohort study, we revealed first time in literature the gender differences in admission to hospital and rates of mortality for diabetic patients with COVID-19.

The demographics, length of stay, mortality rates and concomitant chronic metabolic diseases of 152 patients diagnosed with COVID-19 were found in our hospital electronic document system (Probel) and recorded in excel files for further statistical analysis.

In the mortality group (n:22), the numbers of men and women were 9 (40.9%) and 4 (18.2%), respectively. Comparing gender rates in diabetic group, the mortality risk of diabetic men was higher and statistically significant (p<0.05, Pearson Chi-square value:7.246).

We hope that the findings of this research will give scientists an idea of gender differences in viral pandemics for further studies.

Human papillomavirus (HPV) is the fourth most common cause of cervical cancer (CC). The aim of the present study was to investigate gene expression levels of previously identified transcriptional signatures for malignant and non-malignant CC.

To validate of previously analyzed microarray gene expression data, we selected two hub genes (CDK1 and PLK1) and four differentially expressed mRNAs that were common in pre-malignant-normal and malignant-pre-malignant networks (SMS, NNT, UHMK1 and DEPDC1). To this purpose, the study included women diagnosed histologically with malignant CC (n=15), pre-malignant (n=15), and normal subjects (n=15). The expression of six host genes and viral E6/E7 genes were measured by quantitative Real-Time PCR.

The results showed higher expression of CDK1/PLK1 hub genes and SMS, NNT and UHMK1 genes in malignant CC group than non-malignant CC group and normal group. A positive correlation was observed between gene expression of viral E6/E7 oncogenes and UHMK1 gene.

Dysregulation of several mRNA signatures are a common feature of CC and can be potentially used as diagnostic and prognostic biomarkers as well as can be applied to therapeutic targets for CC treatment.

Infections is yet one of the life-threatening complications of the hematopoietic stem cell transplantation (HSCT). The myeloablative and immunosuppressive conditioning regimens, which are administered before HSCT, dampen the defense capacity of the recipients’ immune systems. In this condition, opportunistic infections, especially viral infections such as cytomegalovirus (CMV) can be reactivated and cause morbidity and mortality in HSCT patients. Here, we aimed to find out any possible relationship between types of conditioning regimen and CMV reactivation in allogeneic HSCT patients.

We retrospectively analyzed the data of 145 CMV-seropositive cases out of total 201 allo-HSCT patients, including age, gender, underlying disease, conditioning regimen, prophylaxis regimen and occurrence of acute graft-versus-host disease (aGVHD) to evaluate their roles in CMV reactivation.

Our result showed that conditioning regimen containing Busulfan and Fludarabine (P=0.003) or Cyclophosphamide (P=0.02) significantly decrease the early CMV reactivation. Patients who developed aGVHD (P=0.003) and those who received anti-thymocyte globulin (ATG) as prophylaxis regimen (P=0.002), had 1.84 and 2.63 times higher risks of CMV reactivation, respectively.

Our findings suggest the conditioning regimen, aGVHD and ATG as influencing factors for early CMV reactivation post-HSCT which should be considered in the future studies.

Hepatitis C is the most common hepatotropic viral infection that affects patients on maintenance hemodialysis. Most of the laboratories in India depend on HCV antibody detection by ELISA. PCR based studies on detection of HCV RNA among haemodialysis patients are very scanty in India. The current study was undertaken to find the prevalence of HCV among haemodialysis patients by ELISA and PCR.

This prospective study was conducted from January to May 2018 in a total of 100 samples. Patients more than 18 years of age, who had undergone at least 15 sessions of dialysis were enrolled in the study. All samples were screened for HCV antibody by ELISA and HCV RNA by PCR. Data regarding age and gender of the patients, history of blood transfusion, duration of hemodialysis, total bilirubin levels were collected from medical records.

Among the 100 samples, only one was positive for HCV antibody by ELISA. Eight samples were positive for HCV RNA by PCR. In this study 62.5% of the HCV positives had a previous history of blood transfusion. Duration of dialysis was more among the HCV positive group but there was no statistical significance.

This is the first study from the southern state of Kerala in India showing the prevalence of HCV among hemodialysis patients by PCR. Our study showed an overall HCV prevalence of 8% by PCR. All the PCR positive samples were negative by 3rd generation ELISA which is an alarming finding and further justifies the need for PCR for detecting HCV.

Co-infection of hepatitis C virus (HCV) with human immunodeficiency virus (HIV) is increasing due to similar transmission pathways. Chronic HCV infection is the most common complication among HIV-infected individuals. Information on the frequency of HCV infection on Iranian HIV-infected individuals is scarce. The aim of this study was the detection of HCV prevalence and genotypes among HIV-infected people in Sanandaj, Iran.

In this cross-sectional study, whole blood samples were taken from 185 HIV positive individuals referring to Consultation Center for Behavioral Diseases, Sanandaj, Iran. The ELISA test was done on samples for anti-HCV antibodies. RNA was extracted from only anti-HCV antibody positive samples. An RT-PCR test was conducted to detect HCV RNA. Genotypes of HCV were detected by melting curve analysis by specific primers and probes. Test results and demographic information were analyzed by SPSS software.

The mean age of individuals was 39.3 ± 9.4 years. Out of 185 individuals 99 (53.5%) were positive for anti-HCV antibodies. Out of 99 antibody positive individuals, 44 had HCV RNA. Among 44 RNA positive individuals, genotypes and subtypes of HCV were as 26 (59.1%) 1a, 17 (38.6%) 3a and one (2.2%) 4. There was a significant association between anti-HCV antibody and demographic variables including, age, gender, occupation, and CD4+ T-cell count (p = 0.0001).

The present study reveals that HIV/HCV co-infection is high in the study population. It is recommended similar studies should be done in other HIV infected populations for management of HIV/HCV co-infection.

No abstract