Iranian Journal of Microbiology
Volume:13 Issue: 1, Feb 2021

The magnitude and pace of global affliction caused by Coronavirus Disease-19 (COVID-19) is unprecedented in the recent past. From starting in a busy seafood market in the Chinese city of Wuhan, the virus has spread across the globe in less than a year, infecting over 76 million people and causing death of close to 1.7 million individuals worldwide. As no specific antiviral treatment is currently available, the major strategy in containing the pandemic is focused on early diagnosis and prompt isolation of the infected individuals. Several diagnostic modalities have emerged within a relatively short period, which can be broadly classified into molecular and immunological assays. While the former category is centered around real-time PCR, which is currently considered the gold standard of diagnosis, the latter aims to detect viral antigens or antibodies specific to the viral antigens and is yet to be recommended as a stand-alone diagnostic tool. This review aims to provide an update on the different diagnostic modalities that are currently being used in diagnostic laboratories across the world as well as the upcoming methods and challenges associated with each of them. In a rapidly evolving diagnostic landscape with several testing platforms going through various phases of development and/or regulatory clearance, it is prudent that the clinical community familiarizes itself with the nuances of different testing modalities currently being employed for this condition.

Several studies have focused on the alterations of hematological parameters for a better understanding of the COVID-19 pathogenesis and also their potential for predicting disease prognosis and severity. Although some evidence has indicated the prognostic values of thrombocytopenia, neutrophilia, and lymphopenia, there are conflicting results concerning the leukocyte and monocyte count.

In this retrospective Double Centre study, we reviewed the results of WBC and monocyte counts of 1320 COVID-19 patients (243 of whom (18.4%) had severe disease) both on admission and within a 7-day follow-up.

We found that both the number of monocytes and the percentage of monocytosis were higher in the severe group; however, it was not statistically significant. On the other hand, we found that not only the mean number of WBCs was significantly higher in the severe cases also leukocytosis was a common finding in this group; indicating that an increased number of WBC may probably predict a poor prognosis. Also, the monocyte count was not affected by age; however, univariate analysis showed that the percentage of leukocytosis was significantly greater in the older group (>50) with an odds ratio of 1.71 (P: 0.003).

Alteration of monocytes either on admission or within hospitalization would not provide valuable data about the prediction of COVID-19 prognosis. Although the rapidly evolving nature of COVID-19 is the major limitation of the present study, further investigations in the field of laboratory biomarkers will pave the way to manage patients with severe disease better.

Infection with Infectious bronchitis virus (IBV) and avian pathogenic Escherichia coli (APEC) is an important respiratory infection worldwide. Apoptosis is a physiological process of cell death that occurs as part of normal development and responds to a variety of physiological and pathophysiological stimuli. The identification of molecular mechanisms of action or inaction of key apoptotic proteins is important. This study aimed to investigate apoptotic related genes in the trachea tissue of infected (IBV variant 2, and APEC serotype O78: K80) SPF chickens group compared to the control group.

Forty SPF chickens was divided into 2 groups. Differential transcriptional profile in the infected SPF chickens trachea tissue was compared to those of control group in the early stage of infection by Illumina RNA-seq technique paired-end and strand-specific sequencing. Differentially expressed genes (DEGs) of transcriptome profiling of the trachea from the infected group were identified. Gene ontology category, KEGG pathway, and STRING analysis were analyzed to identify relationships among differentially expressed genes.

Twenty-eight apoptotic genes were identified. They consisted of six pathways related to cell death: the extrinsic pathway, intrinsic pathway, endoplasmic reticulum stress pathway, MAPK signaling pathway, and cell death by NFkB and activates mTOR pathway and some regulator and apoptosis inhibitors.

All of the apoptotic genes in our study were up-regulated. Among these genes, the more fold change value was for TRADD and BCL2A1 genes, and the less fold change value was for MAP3K14, NFKB1, PIK3CB, and ITPR2 genes.

Pertussis is an infectious disease caused by the Gram-negative bacterium Bordetella pertussis. In Peru, actual public health programs indicate that vaccination against B. pertussis must be mandatory and generalized, besides all detected cases must be reported. The objective of this study was to determine the prevalence of B. pertussis among children under five years of age with a presumptive diagnosis of whopping cough in Cajamarca, a region located in northern Peru.

The population of this cross-sectional study were children under 5 years old hospitalized as presumptive cases of pertussis during December 2017 to December 2018. The nasopharyngeal samples were analyzed by real-time PCR for the detection of B. pertussis.

B. pertussis was identified as PCR + in 42.3% of our sample (33/78). The clinical presentation that was observed most frequently includes paroxysmal coughing (97%), difficulty breathing (69.7%), cyanosis (72.7%) and post-tussive emesis (60.6%). Additionally, pneumonia was the most observed complication (33.3%). Four of the patients with PCR+ for B. pertussis presented only lymphocytosis, five only leukocytosis, two patients with decreased leukocytosis and lymphocytes and only one patient with leukopenia and relative lymphocytosis. There was a percentage of 84.8% of unvaccinated children in the PCR+ group. Finally, the mother was the most frequent symptom carrier (18.2%).

In conclusion, in the studied population there is a high rate of PCR+ cases for B. pertussis. Laboratory values may show leukopenia or lymphopenia in patients with pertussis. It is necessary to use appropriate laboratory diagnostic tests in all infants with respiratory symptoms for B. pertussis. Since, the clinical diagnosis overestimates the diagnosis of pertussis.

Resistance to methicillin in methicillin resistant strains of Staphylococcus aureus (MRSA) is due to the presence of mec-A gene ,which encodes a low affinity penicillin binding protein (PBP)-2a or PBP2. Accurate and rapid identification of MRSA in clinical specimens is essential for timely decision on effective treatment. The aim of the study was to compare three different methods for detection of MRSA namely cefoxitin disc diffusion, CHROM agar MRSA and VITEK-2 susceptibility with PCR which is the gold standard reference method and to find the antibiotic susceptibility pattern of these isolates by VITEK-2.

A Total of 100 non-duplicate S. aureus isolates were collected from different clinical samples among both outpatient and inpatients. Detection of MRSA among these isolates was done by cefoxitin disc diffusion, VITEK-2, CHROM agar MRSA and PCR.

The sensitivity and specificity of cefoxitin disc diffusion and Vitek was found to be 97.2% and 100%, while that of CHROM agar was found to be 100% and 78.6%. The overall prevalence of MRSA in our study by PCR was 72%.

Based on the findings in our study, isolates which show cefoxitin zone diameter < 22 mm can be reported as MRSA. However, those isolates which have a zone diameter between 22-24 mm, should ideally be confirmed by PCR.

Pseudomonas aeruginosa is a problematic opportunistic pathogen causing several types of nosocomial infections with a high resistance rate to antibiotics. Production of many virulence factors in P. aeruginosa is regulated by quorum sensing (QS), a cell-to-cell communication mechanism. In this study, we aimed to assess and compare the inhibitory effect of azithromycin (AZM) and EPI- PAβN (efflux pump inhibitor- Phenylalanine-Arginine Beta-Naphthylamide) on QS system and QS-dependent virulence factors in P. aeruginosa clinical isolates.

A total of 50 P. aeruginosa isolates were obtained from different types of clinical specimens. Isolates were investigated for detection of QS system molecules by AHL cross-feeding bioassay and QS-dependent virulence factors; this was also confirmed by detection of QS genes (lasR, lasI, rhlR, and rhlI) using PCR assay. The inhibitory effect of sub-MIC AZM and EPI PAβN on these virulence factors was assessed.

All the P. aeruginosa, producing QS signals C4HSL, failed to produce C4HSL in the presence of sub-MIC AZM, In the presence of EPI PAβN (20 µg/ml) only 14 isolates were affected, there was a significant reduction in QS-dependent virulence factors production (protease, biofilm, rhamnolipid and pyocyanin) in the presence of either 20 µg/ml EPI or sub-MIC of AZM with the inhibitory effect of AZM was more observed than PAβN.

Anti-QS agents like AZM and EPI (PAβN) are useful therapeutic options for P. aeruginosa due to its inhibitory effect on QS-dependent virulence factors production without selective pressure on bacteria growth, so resistance to these agents is less likely to develop.

Bacterial antibiotic resistance is one of the most important threats for public health around the world. Carbapenemase-producing Gram-negative bacteria have resistance to most antibiotics including carbapenems complicating the treatment of infections. The aim of this study was to determine the antimicrobial susceptibility pattern of carbapenemase-producing nosocomial Gram-negative pathogens at a referral teaching hospital to reveal the best options for treatment of related infections.

Gram-negative bacteria, isolated from hospitalized patients with nosocomial infections, underwent meropenem susceptibility test by disk diffusion method. Meropenem-resistant strains were evaluated for the presence of carbapenemase using Modified Hodge test (MHT). Finally, the antibiotic susceptibility test was performed to determine the sensitivity of each carbapenemase-positive strain against various antimicrobial agents according to the guidelines of Clinical and Laboratory Standards Institute (CLSI).

Over the study period, 155 carbapenemase-positive isolates were detected. Pneumonia was the most frequent related nosocomial infection (67.1%) followed by UTI (23.2%). Acinetobacter baumannii (53.5%) and Klebsiella pneumoniae (40%) were the most frequently isolated pathogens. The pathogens had high rate of resistance to all antibiotics. Colistin had the most in vitro effect against all pathogens. Also, K. pneumoniae had a co-trimoxazole sensitivity rate equal to colistin (30.6%).

Carbapenemase-positive Gram-negative bacteria causing nosocomial infections are common in our hospital and have high rate of resistance to most antibiotics. Improvement in the pattern of antibiotic use and infection control measures are necessary to overcome this resistance.

Genetic diversity of Mycobacterium tuberculosis clinical isolates from tuberculosis patients in the multiethnic province of Alborz, Iran was assessed.

A total of 17 isolates in the period of 2012-2013 were collected and subjected to a Multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) consisted of 6 variable numbers of tandem repeats (VNTRs) including ETR-A, ETR-B, ETR-C, ElTR-D, ETR-E, ETR-F, 5 Mycobacterial Interspersed Repetitive Units including MIRU10, MIRU16, MIRU26, MIRU39, MIRU40, and 1 Queen University of Belfast locus, QUB11.

This classified all isolates into 17 distinct MIRU-VNTR types, a reflection of a highly heterogenic population. Within the 12 used VNTR loci, ten proved highly or moderately discriminant according to the calculated HGDI scores. No cluster of isolates was identified in the study panel, giving a clustering rate of 0%, several events of SVL (N=5) and DVL (N=4) and TVL (N=3) were detected.

The greater heterogeneity observed here by MLVA-VNTR analysis is most likely due to limited background data in the study region rather than a genuine more heterogeneous population compared to other provinces of the country.

Staphylococcus aureus is frequently involved in bovine subclinical mastitis worldwide. Besides, the methicillin-resistant S. aureus (MRSA) carrier state of animals is a matter of worrisome. This study aimed to evaluate the frequency of MRSA, discriminatory geno-analysis and antibiotic resistance scheme of the strains isolated from bovine subclinical mastitis in Kurdistan province of Iran.

A total of 283 samples were collected and analyzed for S. aureus phenotypically and molecularly. SCCmec and coa types, and pvl gene were evaluated using polymerase chain reaction (PCR). Finally, the restriction fragment length polymorphism (RFLP) patterns of coa types and the antimicrobial susceptibility profile of the isolates were assessed.

Among the 95 isolates of S. aureus, 11 (11.57%) strains were recognized as MRSA. Six, one, and four SCCmec types represented for IVa, IVc, and V were determined, respectively, among which an individual IVa and V determinant harboured pvl gene. Restriction digestion products of 490 bp, 680 bp, and 730 bp of coa bands were generated. Tobramycin, mupirocin, fusidic acid, clindamycin, and chloramphenicol were the most effective drugs against the MRSA isolates.

The detrimental involvement of S. aureus in bovine subclinical mastitis is proved herein. Besides, the contribution of MRSA and potential contamination of milk and dairy products with the bacterium may impose a serious public health risk. This demands serious and long-lasting efforts to control the infection. The results may be effective in the implementation of accurate controlling strategies.

Escherichia coli and some Salmonella serovars cause various disease manifestations in poultry leading to significant economic losses. The widespread and imprudent use of antibacterial agents in poultry flocks have increased resistant to many antibacterial agents which has become a major public health concern. Some medicinal plants may be alternative to antibacterial agents. The purpose of this study was to investigate the antibacterial and anti-biofilm activity of summer savory essential oil against E. coli and Salmonella isolated from poultry.

The essential oil was extracted using a Clevenger apparatus and subsequently its compounds were determined using GC-MS. Antibacterial properties of essential oil were determined by disc diffusion method, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). To evaluate the anti-biofilm properties the Microtiter plate test was used. Herbal essential oil was extracted and its compounds were identified correctly.

The major components of Satureja hortensis essential oil were thymol (41.28%), γ-terpinene (37.63%), p-cymene (12.2%) and α-terpinene (3.52%). The inhibition zone diameter in the disc diffusion test for E. coli and Salmonella were 32 ± 3 and 38 ± 4 mm, respectively, which was confirmed by MIC and MBC values. Regarding anti-biofilm activity, the MIC/2 concentration of S. hortensis significantly inhibited biofilm formation of E. coli. However, inhibition of biofilm formation of Salmonella was shown at concentration of MIC/2 and MIC/4.

Based on our results, S. hortensis essential oil showed the growth inhibition and bactericidal activity against E. coli and Salmonella. Moreover, this study demonstrated anti-biofilm activity of S. hortensis essential against both tested bacteria.

Tsukamurella species are Gram-positive rods that exist in a broad range of environments. In this study, the efficacy of heat-killed Tsukamurella inchonensis on growth performance, intestinal morphology, and humoral immune responses of broiler chicken was evaluated.

Ross broiler chicks in the cage were randomly allocated to five groups. Trail diets were prepared by adding 106 cells per bird of heat-killed T. inchonensis into the basal trading diet for group 1 continuously dosed for 24 h from day 1 to day 13, and for group 2, 24 h on days 1 to 5; 8; 9, 12 and 13. Group 3 was received 106 bacteria as a subcutaneous injection on days 1, 6, and 12. Groups 4 and 5 were not received T. inchonensis during the experiment period.

Feed intake (FI) and feed conversion ratio (FCR) were not altered by different delivery methods of T. inchonensis supplementation. The pulsed dosed in feed tended to provide higher body weight gain (BWG) than the negative control groups. T. inchonensis treatments, never less of the ways of delivery, boosted (P<0.05) the antibody titers to Newcastle disease virus (NDV), and avian influenza (AI) (H9N2) virus, especially when broiler chickens treated with pulse dosed in the feed. The most significant intestinal development (p<0.05) was observed between groups 1 and 2. There were no significant differences in the thymus, liver, and bursa of Fabricius relative weight. Still, there were significant increases in the relative weight of spleen on day 14 in vaccinated chickens treated with T. inchonensis pulse dosed.

It seems that the supplementation of T. inchonensis in the broiler diet can improve intestinal morphology and humoral immune response, which was represented by increased antibody response to NDV, and AI vaccines significantly, but it cannot affect FI and FCR.

Bacillus probiotics have been recently considered in biotechnological researches, and food additives. The present study was aimed to investigate the effects of Bacillus subtilis probiotics (PY79 and ATCC 6633) and their metabolites on Salmonella Typhimurium in Caco-2 cells.

Cytotoxicity of B. subtilis ATCC 6633 crude supernatant (CS) was evaluated by 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. S. Typhimurium invasion assay was performed in the presence of the probiotics. Cell viability, apoptosis, and necrosis were evaluated in presence of S. Typhimurium , B. subtilis strains, and CS (4%, 8%) using flow cytometry.

Results showed a significant reduction in the invasive ability of S. Typhimurium to Caco-2 cells by employing B. subtilis probiotics, and CS‏) ‏p < 0.05‎‏). The less invasion was indicated in B. subtilis PY79 and Salmonella co-cultural group. Furthermore, the cell survival rates, and apoptosis/necrosis were respectively increased and decreased in co-culture groups (‏p < 0.05‎‏).

Hence, it seems that B. subtilis strains could be suggested as beneficial candidates to overcome the invasion and ‎cytotoxicity of Salmonella on the intestinal cells. However, additional in vivo models are suggested to validate our results.

Haplopappus multifolius Phil. Ex Reiche and Haplopappus taeda Reiche are medicinal shrubs native to Chile and are popularly known as "Bailahuén". Regularly, this plant is used for liver, digestive and renal affections, as well as colds and the cleaning of infected wounds. The aim of the study was to identify the responsible compounds for the antimicrobial activity of H. multifolius and H. taeda.

Infusions and ethanolic extracts of H. taeda and H. multifolius were analysed by thin-layer chromatography bioautography (TLC-B) to determine the compounds responsible for the antimicrobial activity against Gram-positive and Gram-negative bacterial strains and yeasts of Bailahuén. Finally, the minimum inhibitory concentration (MIC) of pure compounds isolated was determinate.

Extract of Bailahuén had activity only against Gram-positive bacterial strains and this activity was associated with aesculetin, 18-acetoxy-cis-cleroda-3,13E-dien-15-oic acid and aromadendrin-7-methyl ether compounds.

H. multifolius and H. taeda have antibacterial capacity on different species of Gram-positive bacteria pathogenic for humans.

Dental caries is one of the most common chronic diseases around the world. Inhibitory effects of Magnolia Grandiflora bark extract has been proved on tooth decay both in vitro and by using free sugar chewing gum. This research aimed to examine the effect of Magnolia Grandiflora bark mouth-wash on the prevalence of Streptococcus mutans in dental plaque.

This crossover, placebo-controlled, clinical trial study, was performed on a total of twenty participants (aged 18 to 35 years) in both control and intervention groups and four phases. The prevalence of S. mutans was measured in a certain volume of volunteer’s dental plaque at the beginning of the project (phase 1), after the first prescription (phase 2), following the washout period (phase 3) and finally after the second prescription (phase 4) by culture on bacteriology medium. Plaque index and saliva sampling were carried out in follow-up visits by a dentist. The data were analyzed using T-Test (paired and independent) quantitatively.

There was a significant difference in S. mutans frequency in dental plaque between when the participants used Magnolia mouthwash and when they washed out or used a placebo (p<0.005). Results also showed a significant difference between Magnolia and Placebo groups in the mean count of saliva bacterial colony counts after oral administration in the first and second time (P<0.001 and P<0.004, respectively).

The current trial showed that Magnolia Grandiflora %0.3 mouthwash tends to decrease the number of S. mutans in dental plaque significantly. Therefore, its mass production and release to the oral health community are suggested. However, further studies with larger sample sizes and varying treatment are required to substantiate the findings of this study.

Acinetobacter baumannii is recognized as an important pathogen responsible for serious infections causing episodes of hospital infection. Carbon nanotubes (CNTs) have recently emerged as superior materials against antibiotic-resistant bacteria. In this study, a new chemical compound was designed in order to combat A. baumannii infections. Subsequently, the effect of this novel carbon nanotube coated with an antibacterial compound on Extensively Drug-Resistant (XDR), Multidrug-Resistant (MDR) and Pan-Drug-Resistance (PDR) strains of A. baumannii was investigated.

A total of 122 clinical isolates of A. baumannii were cultured from burn patients and their susceptibility to antibiotics were checked using disk diffusion method and Minimum inhibitory concentration. Antimicrobial effects of the coated carbon nanotube were evaluated on XDR, MDR and PDR isolates of A. baumannii. Cell viability was determined using tetrazolium reduction assay (MTT) on human fibroblast cell line (HDFa). Wound healing processes were assessed by quantitative polymerase chain reaction.

Of the 50 A. baumannii isolates, 38 (76%) were found to be MDR and 12 (24%) were XDR. No PDR strains were detected. Results indicated that the carbon nanotube combined with mercury had antibacterial effect against different A. baumannii species and it also was able to increase the expression of epidermal growth factor, platelet-derived growth factor and vascular endothelial growth factor A mRNA levels which are involved in wound healing.

The engineered carbon nanotube compound can potentially be used for treatment of burn related infections. This can potentially give clinicians a new tool for treating A. baumannii infections.

Plant Growth-promoting Bacteria (PGPB) can replace the dangerous chemical fertilizers and pesticides. The aim of this study was to isolate the PGPBs for Lycopersicon esculentum plant and to determine the appropriate volume for inoculation.

Plants samples were collected from tomato fields. Nitrogen fixing-PGPBs were isolated from rhizoplane and rhizosphere. Five isolates were screened based on their growth abilities and examined for PGPB traits including phosphate solubilization, and IAA, ammonia and HCN production. After high cell density cultivation, the cells were separated by centrifugation and freeze dried after resuspension in cryoprotectant. The powders were inoculated into sterile soil with a dose of 106, 107 and 108 CFUs/g. Tomato (Lycopersicon esculentum) seeds were sown in soil and after 42 days the shoot length was measured.

Most of the potent PGPBs with high growth capacity were isolated from rhizoplane. Maximum phosphate solubilization was 289.7 µg/ml by NFB12 which isolated from rhizoplane. This strain produced the maximum level of IAA. NFB12 produced ammonia without the ability of production of HCN. This strain enhanced shoot length in dosed dependent manner. Surprisingly, inoculation of soil with 108 CFUs/g dramatically decreased the shoot length by 21%. Based on molecular approach NFB12 was identified as Bacillus megaterium.

Isolation of specific PGPBS is recommended for sustainable plant production. Our results showed that NBF12 improves tomato plant growth and its effect on tomato plant growth is does dependent. Maximum growth rate of tomato was observed with 107 CFUs/g soil inoculation of NFB12 while higher inoculation showed negative effect.

This study was aimed to isolate Rhizobium spp., from the plant rhizosphere and to investigate their effects on the growth of peanut (Arachis hypogaea L.) as plant growth-promoting rhizobacteria (PGPR).

The isolates were characterized using YEMA, YEMA + Congo Red, and YEMA + Bromothymol blue (BTB) media. The Rhizobium was tested qualitatively for their ability to produce indole acetic acid (IAA), siderophores, proteases, nitrogenases as well as phosphate solubilizing activity. A greenhouse experiment was carried out to elucidate the effect of Rhizobium inoculation on Arachis hypogaea L. growth.

Eleven isolates were obtained in YEMA media and they were red-pink in the YEMA + Congo Red media. The YEMA + BTB test showed that 2 isolates were slow-growing and the rest were fast-growing isolates. Seven isolates produced siderophores, 5 were capable of phosphate solubilizing, 9 isolates produced protease enzyme, 4 isolates could produce IAA, and 7 isolates could fix nitrogen. The B1 and the combination of some high trait-isolate treatments in Y gave the best results on Arachis hypogaea L. growth.

These isolates can be developed as biological fertilizer agents for the peanut plant.

The three old world Leishmania species i.e., L. major, L. tropica, and L. infantum are considered as potential etiological agents of the various clinical forms of leishmaniasis in Iran. Different species co-exist in some areas. Accurate differentiation between the species is essential for choosing an appropriate therapy. Conventional and gold standard methods for the detection and characterization of parasites are time-consuming, laborious, and have low sensitivity. A polymerase chain reaction followed by high resolution melting (PCR-HRM) analysis has been employed for detection and species identification. Most of the studies suffer from the use of multiple targets and/ or requiring more than one reaction to identify a single sample. The present study aimed to design a PCR method based on the amplification of kinetoplast DNA minicircles (kDNA) and HRM analysis of the amplicons for rapid discrimination of the three mentioned species.

DNA from reference strains including L. major, L. tropica, and L. infantum and fifty-eight strains subjected to PCR-HRM analysis targeting kDNA. All the samples were also analyzed by conventional kDNA- PCR.

The PCR-HRM analysis allowed discrimination between the three Old World species. The normalized HRM curves for the amplicons of kDNA indicated a unique and repeatable melting plot for each species, even in combination with human and mouse genomic DNA. Conventional kDNA- PCR could not properly discriminate L. tropica from L. infantum.

PCR- HRM analysis of kDNA proved to be fast and accurate for discrimination of L. major, L. tropica, and L. infantum.