医学微生物学与诊断

Pathogenic Staphylococcus aureus is the most frequently isolated Gram positive bacterium from clinical specimens; and it is among the leading cause of infection in man. S. aureus has gained significant interest in recent years as an important nosocomial pathogen – owing to its multidrug resistant nature which is associated to several virulence factors of the organism including Panton-valentine leukocidin (PVL). PVL is part of the toxins produced by pathogenic S. aureus – which help the organism to exacerbate their pathogenicity/virulence in the phase of an infection. This study evaluated the prevalence of PVL-positive S. aureus from clinical specimens – owing to the dearth of information on this subject matter in Nigeria. Out of the 118 non-consecutive S. aureus isolates employed for this study, only 56 isolates were biochemically confirmed as pathogenic S. aureus. The antibiogram showed that the S. aureus isolates were most susceptible to gentamicin, vancomycin, ciprofloxacin, erythromycin and linezolid. However, they were highly resistant to the cephamycin, cefoxitin (82.1%). The S. aureus isolates also showed reduced susceptibility to tigecycline (71.4%), clindamycin (66.1%), chloramphenicol (48.2%), amoxicillin-clavulanic acid (53.6%) and sulphamethoxazole-trimethoprim (48.2%). The prevalence of PVL genes in this study was 10.7%. Only 6 isolates of S. aureus (10.7%) were confirmed by PCR to harbour the PVL genes; and these S. aureus isolates were from wound samples, abscess and urine samples. The occurrence of pathogenic S. aureus harbouring drug resistant genes such as PVL genes in the hospital environment pose serious health and therapeutic challenges especially in choosing antimicrobial therapy for treatment. S. aureus isolates with PVL genes could also disseminate with high propensity within the hospital environment; and this could result in the outbreak of nosocomial infections. Continues antibiotic stewardship in our hospitals will help in the control and prevention of the emergence and dissemination of drug-resistant microbes in both the community and hospital environment.

Hepatitis C virus (HCV) infection represents a great public healthcare challenge as it affects nearly 170 million individuals worldwide. Therefore, the deep investigation of the mechanisms involved in the pathogenesis of chronic hepatitis induced by HCV is a crucial step in the design of novel targeted therapies for the treatment of this condition. However, techniques of molecular biology to characterize HCV proteins can suffer of intrinsic limitations due to high mutation rates of the virus genome.

In this study, we propose a novel strategy to synthesize a viral cDNA sequence corresponding to the p7 gene in HCV genome-free conditions. Our approach consists of a three-steps polymerase chain reactions (PCRs) by using a set of four large overlapping synthetic oligonucleotides aimed to separately amplify both 5’ and 3’ ends of the p7 gene; 5’ and 3’ products, overlapping themselves, were then used as a template in a third PCR amplification in order to get a full-length p7 cDNA.

Our methodology represents an interesting proof-of-principle as it allows for the safe manipulation of short viral genes. Moreover, this new technique overcomes the elevated genetic variability of HCV genomes without affecting the antigenic characteristics of the putative viral protein.

Over the past twenty years the worldwide clinically impact of Acinetobacter baumannii (A. baumannii) demonstrated its etiopathogenetic relevance. During a previously retrospective study in a teaching hospital, between January 2011 and February 2015, we observed increasingly infections caused by A. baumannii associated with antibiotic multi-resistance. Tigecycline, the first member of the glycylcycline class, is an effective option for the treatment of such infections even if, due to its increased clinical use, tigecycline resistant isolates have recently emerged. In A. baumannii several mechanisms are associated with a tigecycline decrease susceptibility, among these, expression efflux pump AdeABC and the presence of insertion sequence (IS) in the adeRS operon.About that, we decided to analyze adeB and adeS genes in 24 MDR A. baumannii clinical isolates, selected on the different tigecycline phenotype. The study of adeB and adeS genes was performed by an in-housepolymerase chain reaction (PCR) and by Sanger sequencing method. According to literature adeB and adeS genes were detected in all MDR A. baumannii isolates tested. Therefore our attention has focused on two resistant tigecycline clinical strains (ACI 2313 and ACI 1213), with a MIC value >8. In particular the ACI 2313 strains, showed the presence of an IS in the adeS gene. Then, adeS sequence analysis identified ISAba1 insertion. Moreover, adeB gene expression was evaluated by an in-house SYBR Green I-based real-time RT-PCR. We found an over expression of adeB gene in ACI 2313 strain, according to IS presence on adeS gene, while the lack of adeB overexpression in ACI 1213, still resistant to tigecycline, could be due to different resistance mechanisms.

Abstract

In ancient and modern epoch, aerial parts of herbal plants have been broadly used for the treatment of primary health care and variety of ailment across the world depends on geographical cultivation. Now a day’s researcher focuses their attention to explore the plants having broad spectrum therapeutic activity. On the basis of medicinal plant activity surveillance, the core goal step of current study is to evaluate the susceptible antimicrobial activity of crude 50% hydro alcoholic extract of leaves of different folk medicinal plant i.e Ocimum basilicum L., Cymbopogon citratus (DC.) Stapf., Olea europaea L., Eucalyptus camaldulensis Dehnh. against different clinical isolates (Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae, Bacillus subtilis, Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcu epidermidis, Salmonella typhi and Candida albican) of microbial disease so as to trip up on the other alternatives and overcome the upcoming era of increasing microbial resistance. These isolates were collected from different hospitals and pathological laboratories of Karachi, Pakistan. Extract were obtain by soaking the leaves in 50% methanol and then vacuum dried through Rotavapor while the antimicrobial activity were evaluated by well diffusion method. Overall outcome of current study endorse that among selected herbs, Olea europea possessed broad spectrum antimicrobial activity. Further investigation is needed to develop formulation from the same plant. It’s timely need to explore the antimicrobial activity of other herbs also.

Information entropy (H) and predicted B cell epitope score (Bepipred) were determined for the envelope E protein of Zika viruses (ZIKV) isolated from infected humans and Aedes mosquitos with the aim of identifying E protein regions that may be useful as immunological targets of anti-ZIKV vaccines. Total H of mosquito origin E proteins was 4.2380 greater than that of E proteins of human origin, suggestive of constraints on ZIKV mutation in the human host. Seven invariant peptides (H=0.0) of length 10 amino acids, or greater, were identified. These peptide sequences where H=0.0 were screened for predicted epitopes. The seven invariant peptides were comprised of 93 amino acid residues, 31 of which demonstrated predicted B-cell epitopic activity. The predicted epitopic residues were distributed predominantly to 5 of the 7 invariant peptides. It is proposed that these 5 invariant (H=0) peptides in the E proteins of both human and Aedes mosquito ZIKV represent domains with constrained mutational/evolutionary potential and that epitopes predicted to reside in such invariant domains thus may be stable immunological targets for development of an anti-ZIKV vaccine.

Antibiotics are widely used for treatment of bacterial infections and for prophylaxis during instrumental procedures and in certain conditions such as immunodeficiency and splenectomy. Hypersensitivity (allergic) reactions are unpredictable and can occur in some patients even if they have taken the antibiotic in the past with no reaction. Drug allergy accounts for 11.3% of all adverse drug reactions. Drug allergy drugs can be generally classified (according to the World Allergy Organization) based on timing of symptoms into immediate (Immunoglobulin E (IgE) mediated) occurring within 1 hour and delayed (non IgE mediated) allergic reactions occurring after 1 hour. Many patients are mislabeled with drug allergy especially when the diagnosis is made based on history alone. In such cases, a referral to an allergist is important to confirm or exclude allergy through a detailed clinical history, in vitro and/or in vivo testing, as over diagnosis of drug allergy leads to the unnecessary use of broader spectrum and expensive antibiotics contributing to the emergence of multidrug resistant pathogens. Also, in cases of confirmed drug allergy it is important to establish potential cross reactivity with other drugs. Equally, patients with confirmed drug allergy, who have an absolute requirement for the drug or cross reactive drug (as in penicillin allergic females with syphilis) can undergo a process of desensitization in order to complete their treatment through induction of temporary tolerance of the drug.

In the present study, the water extracts of Averrhoa bilimbi at different maturity stages were evaluated to investigate antimicrobial activity against two Gram positive and three Gram negative bacteria by disc diffusion and broth dilution assays. All of the bacterial isolates showed varying degrees of sensitivity towards A. bilimbi extracts. For disc diffusion assay, Gram positive bacterium, Staphylococcus aureus was more sensitive to the extract than Bacillus cereus with inhibition zone of 9.3 mm (young fruit), 12.3 mm (mature fruit) and 10 mm (ripe fruit). The findings also demonstrated that the extracts have stronger antimicrobial effects against Gram negative bacteria, Salmonella spp. with inhibition zone of 12 mm at young fruit, 11 mm at mature fruit and 9.3 mm at ripe fruit than Escherichia coli and Pseudomonas aeruginosa. From broth dilution method, the MIC of extracts were 0.125 gml-1 at young stage, 0.25 gml-1 at mature stage and 0.5 gml-1 at ripe stage against Escherichia coli while 0.25 gml-1 at either young or ripe stage and 0.125 gml-1 at mature stage for S. aureus. The results suggested that the antimicrobial properties are influenced by the maturity stages of the fruit especially at early stage of maturity.