Iranian Journal of Microbiology
Volume:12 Issue: 2, Apr 2020

Despite widespread vaccination programs against pertussis, there has been a worldwide resurgence of the disease in recent years. We aimed to investigate protein composition of outer membrane vesicles (OMV) of Bordetella pertussis (Bp) and to evaluate the immunogenicity of OMV antigens both in the vaccine and the dominant wild type strains in Iran.

The OMV were purified from both vaccine and wild type strains. The immunoreactivity of the OMVs was investigated by exposing sera taken from the patients and the vaccinated infants. The protein profiles of OMVs were compared using two-dimensional electrophoresis. The LC-MS/MS was used to analyse and identify differentially expressed protein spots.

The two type strains showed differences in their 2D gel protein profile. Further analysis of selected proteins from the dominant Iranian strains using LC-MS/MS demonstrated that the identified proteins fell into different functional categories including (i) metabolism, (ii) membrane transport and secretion system, (iii) biosynthesis and degradation, (iv) adaption, adhesion, pathogenicity, conserved hypothetical and protection responses. Moreover, a number of immunogenic proteins were identified including Bp 2434 (serine protease) and Bp 1616 (putative DNA binding protein) from the vaccine and the wild type strains, respectively which could be considered as potential antigens for an OMV vaccine.

OMV Bp could be considered as an alternative vaccine against pertussis, containing the bacterium’s protein antigens that can confer equal efficacy compared to a whole bacterial cell vaccine with advantages such as less side effects and lower costs than acellular pertussis vaccines.

Neonatal meningitis is one of the most important and serious neonatal infections with a high mortality and morbidity rate. The present study aimed to investigate the causes, clinical signs, laboratory parameters and mortality rates in newborns with bacterial meningitis.

This cross-sectional study was performed on 468 neonates aged 2-28 days admitted to NICU in Ghaem Hospital Mashhad, Iran by available sampling method during 2009-2018. Meningitis was confirmed according to positive results of CSF culture and clinical feature. By using researcher-made questionnaire, neonate's individual data including cardiopulmonary resuscitation, the Apgar score of the first and fifth minutes, gestational age, birth weight, clinical symptoms and laboratory data such as ESR, WBC and positive culture of CSF were studied.

Among 468 newborn suspected to infection, lumbar Puncture (LP) was performed for 233 cases (50%). Of 233 neonates, 148 neonates (63.5%) had negative results for CSF culture and 85 cases (36.5%) had positive CSF culture. 94% of cases with meningitis were born premature. Blood culture had positive results in 80% of infants with late-onset meningitis and negative in 20%. The most common clinical findings were respiratory symptoms (94%). Klebsiella pneumoniae and Entrobacter aerugenes were the most common microorganisms of meningitis. Gestational disorders were observed in 55.3% of newborns with meningitis. C-Reactive Protein (CRP) of neonates with meningitis was twice higher than normal cases, and leukocytes and proteins in the CSF in neonates with meningitis were higher than healthy ones. Finally, 36% of neonates with meningitis died in our study. For analyzing the relationships between variables, independent t-test was used after controlling the normality, and Chi-square was used for analyzing the relationship of variables with nominal scale.

The most common pathogens of meningitis were Klebsiella pneumoniae and Enterobacter aerogenes. Respiratory symptoms were the most common clinical signs, and laboratory symptoms included increased CRP, increased leukocytes and proteins in CSF.

Trend analysis reveals that Klebsiella pneumoniae has witnessed a steep enhancement in the antibiotic resistance and virulence over the last few decades. The present investigation aimed at a comprehensive approach investigating antibiotic susceptibility including, extended spectrum beta-lactamase (ESBL) and AmpC β-lactamase (AmpC) resistance and the prevalence of virulence genes among the K. pneumoniae isolates.

Sixty-one K. pneumoniae isolates were obtained from various clinical infections. Antimicrobial susceptibility was performed by disk diffusion method. The Mast® D68C test detected the presence of ESBLs and AmpCs phenotypically, and later presence of ESBL and AmpC genes was observed by polymerase chain reaction (PCR). Multiplex-PCR was performed to investigate various virulence genes.

Amongst 61 K. pneumoniae isolates, 59% were observed as ESBL and 14.7% as AmpC producers. All ESBL producers were positive for blaCTX-M-15, while blaCTX-M-14 was observed in 54.1% isolates. The frequency of AmpC genes was as follows: blaCMY-2 (60.7%) and blaDHA-1 (34.4%). The most frequent virulence genes were those encoding enterobactin and lipopolysaccharide. Presence of mrkD was associated with blaDHA-1 gene, while blaCMY-2 significantly (p≤0.05) correlated with the presence of iutA and rmpA virulence genes. blaDHA-1 positive isolates had urine as a significant source, while blaCMY-2 positive isolates were mainly collected from wound exudates (p≤0.05).

Our results highlight that ESBL and AmpC production along with a plethora of virulence trait on K. pneumoniae should be adequately considered to assess its pathogenesis and antibiotic resistance.

Acinetobacter baumannii has been known as a major pathogen causing nosocomial infections. The aim of this study was to develop multiplex PCR for rapid and simultaneous detection of metallo-β-lactamase (MBL) genes in clinical isolates of A. baumannii.

In this study, we used three sets of primers to amplify the MBL genes including blaOXA-48, blaOXA-23 and blaNDM. The multiplex PCR assay was optimized for rapid and simultaneous detection of MBL genes in A. baumannii strains recovered from clinical samples.

A. baumannii strains recovered from clinical samples were subjected to the study. The multiplex PCR produced 3 bands of 501 bp for blaOXA-23, 744 bp for blaOXA-48 and 623 bp for blaNDM genes. In addition to, no any cross-reactivity was observed in multiplex PCR.

Based on obtained data, the multiplex PCR had a good specificity without any cross reactivity and it appears that the multiplex PCR is reliable assay for simultaneous detection of MBL genes in A. baumannii strains.

There are different sporicidal standard tests with various specifications to deal with products that are claimed for sporicidal activity. The aim of this study was to compare the 7% H2O2 sporicidal efficacy against Bacillus subtilis spores using different standard test methods.

The 7% H2O2 sporicidal efficacy against Bacillus subtilis spores was determined according to the AOAC MB-15-04 standard of carrier test and two standard suspension tests (BS EN 13704, AFNOR NF 72-230) in both clean and dirty conditions and by using different interfering substances including bovine serum albumin, yeast extract and skimmed milk.

The results of suspension tests with 3 × 105 and 2 × 107 CFU/ml of B. subtilis spore concentration demonstrated that the higher spore counts lead to lower efficacy of 7% H2O2. Also, the sporicidal activity of 7% H2O2 was reduced in the presence of interfering substances. Bovine serum albumin, yeast, and skimmed milk showed similar interfering effects in suspension test with 3 × 105 CFU/ml. While, in suspension tests with higher initial spore count (2 × 107 CFU/ml) severity of interfering effects were intensified and distinct. Our results indicated that the carrier sporicidal test in comparison with suspension tests required more contact time to kill B. subtilis spores.

The results of this study showed that it is reasonable to use interfering substances and inoculated carriers in accordance with actual conditions of product usage in a sporicidal test. Interfering substances may reduce the contact surface between H2O2 and test spores; therefore, the sporicidal efficacy of H2O2 was diminished. So applying suspension test in clean condition to verify the claim of sporicidal activity is strongly discouraged.

In recent years, active packaging has been introduced as a new method to better preserve food. Chitosan and nanoclay have been used for preparation of an active nanocomposite with respect to their antimicrobial properties to investigate its effects on the microbial limitation in Gouda cheese.

Nanoclay film, chitosan film, chitosan-based nanocomposites and nanoclay-based nanocomposites were prepared and their antimicrobial properties were evaluated to the microbial limitations of Gouda cheese consist of coliforms, Escherichia coli, Salmonella spp., coagulase-positive Staphylococcus, mold and yeast by agar diffusion method.

The results indicated, the best antimicrobial effect belonged to nanocomposite film with the composition of chitosan 3 wt% by adding nanoclay 1 wt%, which can prevent microbial characteristics of Gouda cheese.

The chitosan and nanoclay nanocomposite had excellent antibacterial activity and performed well against microbial limitations (coliforms, E. coli, Salmonella spp., coagulase-positive Staphylococcus, mold and yeast) of Gouda cheese. Therefore, the nanocomposite may be possibly used as a surface coating in addition to Gouda cheese as well as similar cheeses and other food to enhance microbial characteristics and extend shelf life.

Endophytic actinomycetes have been known as a promising source for new antibiotics discovery against susceptible and resistant forms of pathogenic microorganisms. This study was aimed at determining antibacterial compound from Streptomyces sp. strain B-92 isolated from a medicinal plant Neesia altissima.

Streptomyces sp. strain UICC B-92 was endophytic actinomycetes of N. altissima that obtained from Universitas Indonesia Culture Collection (UICC). Isolation and determination of bioactive compound were carried out using thin layer chromatography (TLC), nuclear magnetic resonance spectroscopy (NMR), and liquid chromatography mass spectrometry (LC-MS) analyses. An in vitro antibacterial assay of pure bioactive compound from the endophytic actinomycetes strain was performed against Bacillus cereus strain ATCC 10876, Escherichia coli strain ATCC 25922, Salmonella typhimurium strain ATCC 25241, Shigella flexneri strain ATCC 12022 and Staphylococcus aureus strain ATCC 25923.

The bioactive compound was identified as 4-((3S,4R,5S)-3,4,5-trihydroxy-6-(hydroxymethyl) tetrahydro-2H-pyran-2-yloxy) phenazine-1-carboxylic acid. In vitro antimicrobial assay showed that bioactive compound of Streptomyces sp. strain UICC B-92 exhibited antagonistic activities against two Gram-positive bacteria, viz, B. cereus strain ATCC 10876 and S. aureus strain ATCC 25923.

The findings of this research showed that, bioactive compound of Streptomyces sp. strain UICC B-92 is suggested a new compound based on glycoside structure and its position.

Excess use of pesticides in agricultural field not only compromised soil fertility but also posed serious threat to water bodies and life in the surrounding environment. The leftover pesticide residue needs to be remediated effectively. Compared to physical, chemical and enzymatic remediation options the microbial remediation is more practical and sustainable.

The Pseudomonas stutzeri smk strain was found to use dichlorvos as the solitary carbon source. Minimal medium supplemented with dichlorvos was used to test ability of bacterium to degrade pesticide aerobically. The metabolites produced by the bacterium were studied with UV-Vis spectrophotometry, HPLC, FTIR and GC-MS techniques. The toxicity studies of neat dichlorvos and P. stutzeri smk degraded metabolites were studied by subcutaneous injection in Mus musculus.

The P. stutzeri smk strain was found to degrade as high as 80% of dichlorvos on 7th day of incubation, at 30 °C temperature and at pH 7. In five steps complete aerobic degradation of 2,2dicholorvinyl dimethyl phosphate (dichlorvos) resulted in production of free methyl and phosphate. The degradation intermediates produced are 2-Chlorovinyl dimethyl phosphate, vinyl dimethyl phosphate, dimethyl phosphate, methylphosphate and finally free phosphate. The histopathological analysis of liver, spleen and thymus of M. musculus were performed to study toxicity of dichlorvos and degraded metabolites.

P. stutzeri smk could result highest aerobic degradation of dichlorvos to produce free methyl and phosphate. Degradation metabolites could reverse largely toxic effects of dichlorvos when studied in M. musculus.

Use of antibiotics as growth promoters in animal feeds has been restricted due to the residues in poultry products such as egg and meat, furthermore to the antibiotic resistant of pathogenic bacteria. The prohibition of their use opens the opportunity for the use of non-antibiotic feed additives such as probiotics. The objectives of this study were to investigate the effect of the addition of Lactobacillus casei WB 315 and crude fish oil (CFO) to diets on growth performance, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), low density lipoproteins (LDL), high density lipoprotein (HDL), and cholesterol levesl of broiler chickens.

In this research, one-day old male broiler chicks were used and divided equally into four groups, namely a basal diet without L. casei WB 315 and without CFO (P0), basal diet supplemented with 0.5% L. casei WB 315 of total broiler basal feed (1.2 × 109 cfu/ml) and without CFO (P1), basal diet supplemented without L. casei WB 315 and 1% CFO of total broiler basal feed (P2), and basal diet supplemented with 0.5% L. casei WB 315 of total broiler basal feed (1.2 × 109 cfu/ml) and 1% CFO of total broiler basal feed (P3) for 35 days.

The results of addition 0.5% Lactobacillus casei WB 315 (1.2 × 109 cfu/ml) and 1% CFO of total broiler basal feed after 35 days showed significant difference among treatment in feed efficiency (p<0.05), feed conversion ratio (p<0.05), feed consumption (p<0.05), EPA (p<0.05), DHA (p<0.05), increase HDL (p<0.05), reduced the LDL (p<0.05), and reduce cholesterol (p<0.05) in meat broiler chicken.

It is concluded that the addition of L. casei WB 315 and crude fish oil (CFO) could significant improve the growth performance (feed efficiency, feed conversion ratio, feed consumption) and could significantly improve EPA, DHA and increase HDL and decrease LDL in meat poultry product.

Hepatitis C virus and Human Immunodeficiency Virus (HIV) share the same rate of transmission. HIV/HCV co-infected individuals may result in faster progression of liver fibrosis and highly increase the risk of cirrhosis, hepatocellular carcinoma development. Thus this study was conducted to determine co-infection of HCV genotypes in positive HIV patients in Ahvaz city, Iran.

The sera samples were collected from confirmed 78 infected HIV, 67 (85.89%) males and 11 (14.1%) females. All sera samples were tested for HCV Ab using ELISA test. The HCV Ab positive samples were tested for detection of 5' untranslated (UTR) and core regions of HCV genome using nested RT-PCR. The PCR products of 5UTR and core regions were sequenced to determine HCV genotypes.

Among the 78 infected HIV, 25 (32.05%) cases including 20 (25.64%) males and 5 (6.41%) females were positive for HCV Ab (p=0.316). 53 (67.94%) of HIV patients were negative for HCV Ab. Among 25 positive HCV Ab, 19 (24.35%) cases including 15 (19.23%) males and 4 (5.12%) females were positive for HCV RNA (p=0.447). The PCR products of 5 positive samples were randomly sequenced. The results of sequences and alignments showed that the detected HCV genotypes were three 3a and two 1a. The occurrence of genotype HCV 1a was found in one male injecting drug user Injecting Drug User (IDU) and one female. The HCV 3a genotype was detected in the three males IDU.

The results of this survey indicated that 32.05% of HIV patients were positive for HCV Ab, among them 24.35% were positive HCV RNA. HCV genotype 3a was dominant and detected in the three males IDU. Regarding the consequences of HIV/HCV co-infection, it is suggested that HCV RNA detection should be regularly checked in individuals infected with HIV.

Severe acute respiratory infections (SARI) remain an important cause for childhood morbidity worldwide. We designed a research with the objective of finding the frequency of respiratory viruses, particularly WU and KI polyomaviruses (WUPyV & KIPyV), human coronaviruses (HCoVs), human respiratory syncytial virus (HRSV) and human parechovirus (HPeV) in hospitalized children who were influenza negative.

Throat swabs were collected from children younger than 5 years who have been hospitalized for SARI and screened for WUPyV, KIPyV, HCoVs, HRSV and HPeV using Real time PCR.

A viral pathogen was identified in 23 (11.16%) of 206 hospitalized children with SARI. The rate of virus detection was considerably greater in infants <12 months (78.2%) than in older children (21.8%). The most frequently detected viruses were HCoVs with 7.76% of positive cases followed by KIPyV (2%) and WUPyV (1.5%). No HPeV and HRSV were detected in this study.

This research shown respiratory viruses as causes of childhood acute respiratory infections, while as most of mentioned viruses usually causes mild respiratory diseases, their frequency might be higher in outpatient children. Meanwhile as HRSV is really sensitive to inactivation due to environmental situations and its genome maybe degraded, then for future studies, we need to use fresh samples for HRSV detection. These findings addressed a need for more studies on viral respiratory tract infections to help public health.

Luliconazole is currently confirmed for the topical therapy of dermatophytosis. Moreover, it is found that luliconazole has in vitro activity against some molds and yeast species. The aim of the present study was to evaluate the efficacy of luliconazole in comparison to routine used antifungals on clinical and environmental isolates of Aspergillus flavus.

Thirty eight isolates of A. flavus (18 environmental and 20 clinical isolates) were detected based on morphological and microscopic features and also PCR-sequencing of β-tubulin ribosomal DNA gene. All the isolates were tested against luliconazole, voriconazole, amphotericin B and caspofungin. Minimum inhibitory concentration (MIC), MIC50, MIC90 and MIC Geometric (GM) were calculated using CLSI M38-A2 protocol for both environmental and clinical isolates.

Luliconazole with extremely low MIC range, 0.00049-0.00781 μg/mL and MICGM 0.00288 μg/mL showed very strong activity against both clinical and environmental A. flavus isolates. Moreover, voriconazole inhibited 100% of isolates at defined epidemiological cutoff values (ECV ≤ 2 µg/ml). 50% and 27.8% of clinical and environmental isolates of A. flavus, were resistant to caspofungin, respectively. Whereas, all the isolates were found to be resistant to amphotericin B.

The analysis of our data clearly indicated that luliconazole (with MICGM 0.00244 µg/ml for clinical and 0.00336 μg/ml for environmental isolates) had the highest in vitro activity against A. flavus strains.