Jundishapur Journal of Microbiology
Volume:14 Issue: 1, Jan 2021

Dengue virus (DENV) is an RNA virus belonging to the family Flaviviridae of the genus Flavivirus with worldwide distribution. Dengue fever is caused by any of four closely-related serotypes DENV, an emerging pandemic-prone viral disease in many regions of the world.

The present study aimed to determine the prevalence of dengue virus genotypes and serotypes in children aged below 15 years in Lahore, Pakistan.

In this study, 112 serum samples were collected from clinically suspected dengue fever patients from March 2017 to December 2018 at different tertiary care hospitals in Lahore, Pakistan. Regarding the patients’ age, the samples were divided into four groups from A to D (i.e., 0 - 1, 1 - 5, 5 - 10, and 10 - 15 years of age). Rapid immuno-chromatography (ICT) test was conducted on the collected serum samples, followed by quantitative RT-PCR for serotype of dengue virus.

Out of 112 samples, 34 samples were diagnosed as DENV positive by the rapid ICT screening method. No virus was detected in groups A and B, while three samples were positive in group C (1 boy and two girls), and 31 samples (23 boys and 8 girls) were positive in group D. The results of quantitative RT-PCR exclusively showed DEN-3 serotype in all the ICT positive samples. The results indicated that the prevalence of DEN-3 serotype in children was 100%, indicating that DEN-3 serotype might cause severe epidemics in the future in Lahore, Pakistan. Hematological analysis revealed an increase in hematocrits in 41.1% dengue-positive cases. Leucopenia was prominent in 79.4% of the cases, while Thrombocytopenia was reported in 70.5% of the participants. The biochemical analysis also indicated an increase in liver enzymes in patients (ALT 88%, AST 79%), while the lower levels of cholesterol (69 %) and serum albumin (25%) were also observed.

Dengue virus spreads and grows quickly worldwide over a highly short time interval. Dengue fever claims for a significant number of lives. This study would help individuals know about the status of laboratory parameters in dengue fever and detect how to overcome the prevalence of Dengue virus.

Asthma is a chronic inflammatory disorder of lung airways, affecting about 300 million people worldwide. Several risk factors are involved in asthma development, such as environmental allergens, genetic susceptibility, and respiratory viral infections. Viral infections induce NF-kB and inflammatory pathways that lead to the production of cytokines, chemokines, and inflammatory proteins and, finally, a reduction of lung volume and function.

The aim of this study was to evaluate viral infections’ prevalence in children with asthma from 2016 to 2017.

One hundred throat swab samples were collected from asthmatic children. Extraction of RNA and cDNA synthesis were performed to recognize parainfluenza viruses, rhinoviruses, influenza viruses, and respiratory syncytial virus (RSV) using real-time PCR. Also, the associations of age, sex, and other studied factors with asthmatic attacks were evaluated.

In this study, 41 viruses were detected, including 21 cases of rhinoviruses (51.22%), 10 cases of parainfluenza (24.39%), seven cases of respiratory syncytial virus (17.07%), and three cases of the influenza virus (7.32%). Regarding seasonal incidence, the prevalence of the viruses was high in autumn and winter, and there was a significant relationship between seasonal incidence and gender. However, there were no statistically significant relationships between the prevalence of the viruses and age or gender.

The most important viral causes of childhood asthma in this study were found to be rhinoviruses, followed by parainfluenza. The lowest prevalence was related to the RSV and influenza virus, which the two viruses also showed the lowest seasonal outbreaks. Therefore, it can be said that with an increase in the seasonal incidence of respiratory viruses, the effects of these viruses will be greater on asthma.

Non-tuberculous mycobacteria (NTM) are widely associated with pulmonary diseases. Evidence is lacking on the transmission of NTM from one person to another. Hence, it has gained lower public health priority than tuberculosis.

We determined the prevalence and antibiotic resistance rate of NTM isolated from sputum samples of patients with pulmonary infections.

A total of 375 duplicate sputum samples were collected from 375 patients on consecutive days. The NTM growth was assessed using BACTEC 960 mycobacterial growth indicator tubes. The GenoType Mycobacterium CM/AS line probe assay was used for the species-level identification of mycobacteria. Antibiotic susceptibility tests were performed using the auto-MODS assay.

The overall NTM prevalence rate was 34.4%. Mycobacterium avium complex (24.8%) was the predominant species identified, followed by M. kansasii(24%) and M. abscessus complex (20.2%). Of the 129 NTM isolates tested for antibiotic susceptibility, 62.8% were resistant to rifampicin, 60.5% to levofloxacin, 58.1% to ofloxacin, 55.8% to ethambutol, 49.6% to isoniazid, 48.1% to streptomycin, and 41.9% to amikacin. Seventy-three (56.6%) isolates were identified as multidrug-resistant (MDR) isolates.

Mycobacterium avium complex was the predominant species identified, and the majority of the organisms were resistant to commonly used anti-tuberculosis drugs. The high prevalence of NTM and drug resistance towards the tested antibiotics suggests that NTM can no more be ignored as a contaminant, reiterating the need for periodic surveillance and species-specific treatment for effective management of diseases caused by NTM.

Increasing antibiotic resistance warrants therapeutic alternatives to eradicate resistant bacteria. Combined phageantibiotic therapy is a promising approach for eliminating bacterial infections and limiting the evolution of therapy-resistant diseases.

In the present study, we evaluated the effects of combinations of bacteriophages and antibiotics against multidrugresistant (MDR) Klebsiella pneumoniae.

Two MDR strains (GenBank no. MF953600 & MF953599) of K. pneumoniae were used. Bacteriophages were isolated from hospital sewage samples by employing a double agar overlay assay and identified by transmission electron microscopy. For further characterization of bacteriophages, the killing assay and host range test were performed. To assess therapeutic efficacy, phages (7.5 × 104 PFU/mL) were used in combination with various antibiotics.

The phage-cefepime & tetracycline combinations displayed promising therapeutic effects, restricting the growth of K. pneumoniae isolates, as evidenced by recording OD650nm values.

The results of the current study showed that phage-antibiotic combination was a potential therapeutic approach to treat the infections caused by MDR K. pneumoniae.

The potential of Streptococcus mutans for biofilm formation makes it one of the main organisms causing dental caries. Various preventive strategies have been applied to reduce tooth decay.

In the current study, we aimed to isolate S. mutans bacteriophages from sewage and to investigate their effects on the expression of the genes involved in bacterial biofilm formation in dental caries.

Eighty-one dental plaque samples were collected. Then to isolate and identify S. mutans, bacterial culture media and molecular tests were used. Moreover, the biofilm formation capability of the isolated S. mutans was determined. Also, lytic bacteriophages were isolated from raw urban sewage, and phage morphology was determined by transmission electron microscopy (TEM). Real-time PCR was used to assess the effects of the isolated bacteriophages on the expression of the genes involved in biofilm formation.

Overall, 32 (39.5%) samples were positive for the presence of S. mutans. All of the isolates contained the gtfD gene. The frequencies of other genes were as follows: gtfB (17, 53.12%), gtfC (19, 53.37%), SpaP (13, 40.62%), and luxS (23, 17.87%). The isolated S. mutans bacteria presented different ranges of biofilm formation ability. Based on TEM results, two sewage-isolated bacteriophages, belonging to Siphoviridae and Tectiviridae families, were able to prevent biofilm formation up to 97%.

Our findings indicate that phage therapy can be an optional way for controlling biofilm development and reducing the colonization of teeth surface by S. mutans.

Candida glabrata is the second agent of candiduria with increased resistance to antifungals. Microsatellite length polymorphism (MLP) is one of the genotyping techniques used in the epidemiological investigation to improve clinical management.

We aimed to detect different genotypes of C. glabrata isolates using six microsatellite markers and the MLP technique. Moreover, our genotypes’ association with other countries’ genotypes was illustrated using a minimum spanning tree. We investigated in vitro antifungal susceptibility and enzymatic activity profiles of the isolates.

Six microsatellite markers were amplified using multiplex-PCR for 22 C. glabrata strains isolated from urine in pediatric patients admitted to the Abuzar Children’s Hospital in Ahvaz, Iran. The PCR products were presented for fragment analysis, and the size of the alleles was determined. Antifungal susceptibility tests and extracellular enzyme activities were also performed.

Nineteen multilocus genotypes were detected so that 22.7% of the strains had identical genotypes. The isolates were wildtype for amphotericin B (0.0625 - 2µg/mL), itraconazole (0.125 - 2µg/mL), and voriconazole (0.0078 - 0.00625µg/mL). All the isolates were sensitive to fluconazole at the minimum inhibitory concentration (MIC) range (0.0312 - 16 µg/mL), and three of them were resistant to caspofungin (MIC ≥ 0.5 µg/mL). Moreover, 72.7 and 68.2% of the isolates had no phospholipase and esterase activities. The highest potency of enzymatic activity was obtained in hemolysin and proteinase enzymes. A high genetic diversity (19 genotypes of the 22 isolates) existed among the urinaryC. glabrata isolates. Based on theminimum spanning tree, two clusters of our genotypes were related toC. glabratagenotypes in a previous study in Iran, and the third cluster was entirely connected with Chinese genotypes.

Most of the isolates were the non-wild type for posaconazole but were rarely resistant to other antifungals. Hemolysin and proteinase secreted as the main virulence factors among the urinary C. glabrata isolates.

Leishmania is an intracellular protozoan parasite that uses complex methods for destroying the innate immune response in mammalian host macrophage cells. Many factors have been identified that play a role in the severity of the parasite’s pathogenicity. One of the factors is the GP63, which is a group of metalloproteinases that disrupts the signaling mechanism of the host cell.

The aim of this study was to construct PX-LMGP63 vector through CRISPR-Cas9 for GP63 gene knockout in Leishmania major as a potential method for leishmanization.

A pair of gRNAs were designed based on the mRNA sequence of the GP63. Then annealing primers were cloned into the linearized vector PX-459 and transformed into the DH5A competent cells. Then, PCR assay was performed with gene-specific and vector primers to confirm the colonies. In addition, the constructed plasmid was sequenced for final confirmation.

The expected size band of 270 was confirmed by PCR. The plasmid sequence showed that the gRNA789 was ligated in the vector. The created structure was named PX-LMGP63 and will be transfected into the promastigote cell in the next step.

Owing to the prevalence of cutaneous Leishman as a public health problem in most countries and the lack of an effective vaccine for leishmaniasis, the use of the CRISPR method may make it possible to achieve an effective vaccine in the future.