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Background. Methicillin-resistant Staphylococcus aureus (MRSA) is a pathogenic bacteria involved in a wide spectrum of human diseases. Many virulence factors promote this widespread propagation. One important factor is acquiring antibiotic resistance genes, which leads to a reduction in the availability and efficacy of therapy options. Recently, research has suggested that the remarkable antimicrobial effect of antioxidants against superbugs such as MRSA shows synergistic effects when accompanied by antimicrobial therapy. This paper aims to examine the synergistic effects of ascorbic acid and nicotinamide with a panel of antibiotics used in antimicrobial therapy against MRSA. Material and Methods. Two SCCmec type IV MRSA reference strains (EMRSA-15 and USA300) and 10 MRSA clinical isolates feature in this paper. SCCmec typing was conducted on the 10 clinical isolates via multiplex PCR after identification. Synergy experiments on antioxidants and antibiotics were evaluated via checkerboard assay. The minimum inhibitory concentration (MIC) of each agent was determined in accordance with the Clinical and Laboratory Standards Institute (CLSI) M100 guidelines through twofold microdilution assay. Results and Discussion. Synergy (FIC <0.5) was demonstrated for ascorbic acid (1/2 to 1/4 MIC) with rifampicin (1/2 to 1/8 MIC), and also ascorbic acid (1/2 to 1/16 MIC) when associated with vancomycin (1/2 MIC). Similarly, nicotinamide (1/2 to 1/16 MIC) showed a synergistic effect when paired with low concentrations of rifampicin (1/2 to 1/16 MIC), and also (at 1/4 to 1/16 MIC) with vancomycin (1/2 MIC). All reduced MICs due to synergistic combinations demonstrated statistical significance (P<0.05). Conclusion. The synergistic activity demonstrated in associating antioxidants with antibiotics shows promise in managing superbugs. However, more research is required to better understand the mechanism of the synergy and for utilization in clinical care.

Background. Concurrent coronavirus disease 2019 (COVID-19) and Pneumocystis jirovecii pneumonia (PJP) has been described in various reports, with a recent study describing a 9.3 % P. jirovecii detection rate in critically ill COVID-19 patients. Methods. Patients with PCR-confirmed PJP following COVID-19 infection who were admitted to Aga Khan University Hospital, Karachi, Pakistan from March 2020-June 2021 were identified through a laboratory database. Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus was performed by RT-PCR Cobas SARS-CoV-2 qualitative assay. P. jirovecii PCR was performed using the RealStar Pneumocystis jirovecii PCR kit. Clinical, radiological and laboratory data for PJP patients were recorded. Results. During the study period, 3707 patients were admitted with COVID-19 at our hospital. P. jirovecii PCR was requested for 90 patients and was positive in 10 (11 %). Five out of 10 patients were discharged from the hospital and later developed cough and dyspnoea. Five patients remained hospitalized with severe COVID-19 and developed PJP. Eight patients in our study received systemic steroids. The trends of lymphocyte counts of all patients showed a lymphocyte count of <1000 mm^-3 (<1.0×10⁶ cells µl^-1) in the week of PJP diagnosis. Four patients did not survive; one of these patients did not receive co-trimoxazole due to late diagnosis, one patient had concomitant nosocomial pneumonia and bacteraemia with multidrug-resistant Acinetobacter species, and two patients had concomitant aspergillosis. Conclusion. In summary, invasive fungal infections such as PJP should be considered as a complication in COVID-19 patients, with prompt evaluation and management.

Introduction: Infections forby Myroides spp. can lead to significant morbidity and mortality, particularly in immunocompromised patients with underlying co-morbidities. Recent reports have highlighted its intrinsic and acquired drug resistance, making it a particularly challenging infectious agent to combat.

Methods: Myroides spp. isolated and reported in clinically significant urine samples were considered for the study. Identification of the organism was done via the VITEK 2C system. Antibiotic susceptibility testing was done using both manual and automated methods following Clinical and Laboratory Standards Institute (CLSI) guidelines. Existing literature was searched on MEDLINE using PubMed.

Results: We present a series of five catheter-associated urinary tract infections due to Myroides odoratimimus , with sensitivity to only minocycline. This is the first case from Western India, and the third case in the existing literature that shows Myroides sensitivity only to minocycline. Our literature review is the first to systematically describe contributory factors to infection, allowing us to devise a clinically relevant tool that delineates contributory factors and efficacious drugs in Myroides spp. infection.

Conclusion: Myroides spp. infections, previously considered rare and opportunistic, need cognizance and diagnostic suspicion especially in particular associated conditions.

Francisella tularensis , the causative agent for tularaemia, is a Tier 1 select agent, and a pan-species pathogen of global significance due to its zoonotic potential. Consistent genome characterization of the pathogen is essential to identify novel genes, virulence factors, antimicrobial resistance genes, for studying phylogenetics and other features of interest. This study was conducted to understand the genetic variations among genomes of F. tularensis isolated from two felines and one human source. Pan-genome analysis revealed that 97.7 % of genes were part of the core genome. All three F. tularensis isolates were assigned to sequence type A based on single nucleotide polymorphisms (SNPs) in sdhA. Most of the virulence genes were part of the core genome. An antibiotic resistance gene coding for class A beta-lactamase was detected in all three isolates. Phylogenetic analysis showed that these isolates clustered with other isolates reported from Central and South-Central USA. Assessment of large sets of the F. tularensis genome sequences is essential in understanding pathogen dynamics, geographical distribution and potential zoonotic implications.

Introduction: Shigellosis remains a considerable public health concern in developing countries. Shigella flexneri and Shigella sonnei are prevalent worldwide and S. sonnei has been replacing S. flexneri .

Gap statement: S. flexneri still causes outbreaks of shigellosis in northern Vietnam but limited information is available on its genetic characteristics.

Aim: This study aimed to characterize the genetic characteristics of S. flexneri strains from northern Vietnam.

Methodology: This study used 17 isolates from eight incidents, collected in northern Vietnam between 2012 and 2016. The samples were subjected to whole genome sequencing, molecular serotyping, cluster analysis and identification of antimicrobial resistance genes. Additionally, phylogenetic analysis was performed including isolates from previous studies.

Results: Clusters were identified according to spatiotemporal backgrounds. The results suggested that two incidents in Yen Bai province in 2015 and 2016 were derived from a very recent common ancestor. All isolates belonged to phylogroup (PG) 3, which was divided into two sub-lineages. Thirteen of 17 isolates, including those from the Yen Bai incidents, belonged to sub-lineage Sub-1 and were serotyped as 1a. The remaining four isolates belonged to sub-lineage Sub-2 and were the globally predominant serotype 2a. The Sub-1 S. flexneri isolates possessed the gtrI gene, which encodes the glycosyl transferase that determines serotype 1a, with bacteriophage elements in the vicinity.

Conclusion: This study revealed two PG3 sub-lineages of S. flexneri in northern Vietnam, of which Sub-1 might be specific to the region.

Members of the order Enterobacterales, including Escherichia coli , Klebsiella species and Enterobacter species, are important pathogens in healthcare-associated infections. Higher mortality has been reported from infections due to Klebsiella pneumoniae than from E. coli , but prior studies comparing Enterobacter aerogenes (recently renamed Klebsiella aerogenes ) bacteraemia and Enterobacter cloacae complex bacteraemia have yielded conflicting results regarding whether clinical outcomes differ. We found bacteraemia with K. aerogenes was independently associated with greater risk of 30-day mortality than bacteraemia with Enterobacter cloacae complex.

Giardiasis is an infection of the small intestine caused by the protozoan parasite Giardia intestinalis and one of the most common parasitic intestinal diseases in humans worldwide. It mainly manifests as a self-limited illness in the case of immunocompetent patients and usually does not require treatment. However, immunodeficiency is a risk factor for the onset of severe Giardia infection. In this report, a case of recurrent giardiasis refractory to nitroimidazole therapy is presented. A 7-year-old male patient with steroid-resistant nephrotic syndrome came to our hospital because of chronic diarrhoea. The patient was on long-term immunosuppressive therapy. Microscopic examination of stool showed a significant number of trophozoites and cysts of G. intestinalis. Treatment with metronidazole for longer duration than recommended has failed to clear the parasite in the present case.

Introduction: Patients with coronavirus disease-2019 (COVID-19) are prone to develop respiratory bacterial infections irrespective of their need for mechanical ventilatory support.

Hypothesis/gap statement: Information about the incidence of concomitant respiratory bacterial infections in COVID- 19 patients from India is limited.

Aim: This study aimed to determine the incidence of concomitant respiratory bacterial pathogens and their drug resistance in these patients.

Methodology: A prospective study was performed by including patients who were admitted to our tertiary care centre from March 2021 to May 2021 to evaluate secondary bacterial respiratory co-infections in patients via real-time PCR (RT-PCR)-confirmed cases of COVID-19 disease caused by SARS CoV-2.

Results: Sixty-nine culture-positive respiratory samples from patients with COVID-19 were incorporated into this study. The most commonly isolated bacterial microorganisms were Klebsiella pneumoniae (23 samples, 33.33 %) and Acinetobacter baumannii (15, 21.73 %), followed by Pseudomonas aeruginosa (13, 18.84 %). Among the microorganisms isolated, 41 (59.4 %) were multidrug-resistant (MDR) and nine (13 %) were extensively drug-resistant (XDR). Among the Gram-negative bacteria isolated, K. pneumoniae showed high drug resistance. Fifty carbapenem-resistant microorganisms were isolated from the patients included in our study. Concerning the hospital stay of the patients enrolled, there was an increased length of intensive care unit stay, which was 22.25±15.42 days among patients needing mechanical ventilation in comparison to 5.39±9.57 days in patients on ambient air or low/high-flow oxygen.

Conclusion: COVID-19 patients need increased length of hospitalization and have a high incidence of secondary respiratory bacterial infections and high antimicrobial drug resistance.

The lung microbiome impacts on lung function, making any smoking-induced changes in the lung microbiome potentially significant. The complex co-occurrence and co-avoidance patterns between the bacterial taxa in the lower respiratory tract (LRT) microbiome were explored for a cohort of active (AS), former (FS) and never (NS) smokers. Bronchoalveolar lavages (BALs) were collected from 55 volunteer subjects (9 NS, 24 FS and 22 AS). The LRT microbiome composition was assessed using 16S rRNA amplicon sequencing. Identification of differentially abundant taxa and co-occurrence patterns, discriminant analysis and biomarker inferences were performed. The data show that smoking results in a loss in the diversity of the LRT microbiome, change in the co-occurrence patterns and a weakening of the tight community structure present in healthy microbiomes. The increased abundance of the genus Ralstonia in the lung microbiomes of both former and active smokers is significant. Partial least square discriminant and DESeq2 analyses suggested a compositional difference between the cohorts in the LRT microbiome. The groups were sufficiently distinct from each other to suggest that cessation of smoking may not be sufficient for the lung microbiota to return to a similar composition to that of NS. The linear discriminant analysis effect size (LEfSe) analyses identified several bacterial taxa as potential biomarkers of smoking status. Network-based clustering analysis highlighted different co-occurring and co-avoiding microbial taxa in the three groups. The analysis found a cluster of bacterial taxa that co-occur in smokers and non-smokers alike. The clusters exhibited tighter and more significant associations in NS compared to FS and AS. Higher degree of rivalry between clusters was observed in the AS. The groups were sufficiently distinct from each other to suggest that cessation of smoking may not be sufficient for the lung microbiota to return to a similar composition to that of NS.

Chytrid fungi play key ecological roles in aquatic ecosystems, and some species cause a devastating skin disease in frogs and salamanders. Additionally, chytrids occupy a unique phylogenetic position- sister to the well-studied Dikarya (the group including yeasts, sac fungi, and mushrooms) and related to animals- making chytrids useful for answering important evolutionary questions. Despite their importance, little is known about the basic cell biology of chytrids. A major barrier to understanding chytrid biology has been a lack of genetic tools with which to test molecular hypotheses. Medina and colleagues recently developed a protocol for Agrobacterium -mediated transformation of Spizellomyces punctatus. In this manuscript, we describe the general procedure including planning steps and expected results. We also provide in-depth, step-by-step protocols and video guides for performing the entirety of this transformation procedure on protocols.io (dx.doi.org/10.17504/protocols.io.x54v9dd1pg3e/v1).