The Draft genome sequence of Bacillus subtilis strain S2 Isolated from fermented bamboo drink.
This work reports the draft genome sequence of the fermented food probiotic Bacillus subtilis strain S2 which has antibacterial activity against Vibrio harveyi Staphylococcus aureus Listeria monocytogenes and Streptococcus mutans. From the de novo assembly a chromosome with 4438 open reading frames and an approximate size of 4208679 bp was obtained. Phylogenetically B. subtilis S2 and B. subtilis SRCM103571 are closely related. The organism is known to produce two bacteriocins (uniqueness) Sublancin and Subtilosin A according to investigations done with the use of antiSMASH and BAGEL-4 tools and whole genome sequencing (WGS) analysis. Subsequent analysis showed that this organism's genome with only one complete copy and eight incomplete copies contains the fewest rRNA operons of the ten genomes examined.
ROK or MLCK inhibition restricts Dengue virus replication in endothelial cells
Dengue virus (DV) a member of the Flaviviridae family has been demonstrated to induce changes to the actin cytoskeleton to promote efficient viral entry and replication. These cytoskeletal alterations continue throughout infection and disruption of the cytoskeleton has been shown to decrease the release of infectious virions. However few studies have assessed the contribution of actin alterations to Dengue-induced vascular leak in endothelial cells. A number of cellular kinases control the assembly and rearrangement of actin filaments including Rho-activated kinase (ROK) and Myosin Light Chain kinase (MLCK). To determine whether the observed changes to the actin cytoskeleton during Dengue viral infection contribute to the loss of endothelial cell barrier function the roles of the cellular kinases ROK and MLCK were evaluated during DV infection. Inhibition of ROK and MLCK during infection prevented the formation of DV-induced actin stress fibers. While kinase inhibition did not prevent virus-induced changes in monolayer permeability specific inhibition of either kinase resulted in a significant decrease in virus yield indicating that ROK/MLCK-mediated cytoskeletal alterations play a role in virus replication and biogenesis that do not affect vascular integrity in Dengue virus infection in vitro. Further investigation revealed that ROK or MLCK inhibition decreased virus spread through the culture but only MLCK inhibition decreased viral protein synthesis and genome replication. Taken together these results identify a critical role for both ROK and MLCK activity in the efficient replication of Dengue virus in endothelial cells potentially affecting the assembly or budding of mature
Delving deep into the draft genome of Mangrovibacter sp. SLW1, isolated from Sundarbans mangrove
Mangrovibacter sp. SLW1 a gram-negative aerobic motile bacterium was isolated from mangrove litterfall in Sundarbans mangrove. The draft genome is 5.5 Mbp in size with 49.45 mol% GC content. The linear chromosome of the bacterium consists of 27 contigs with 7339 coding sequences. The genome investigation of Mangrovibacter sp. SLW1 through in depth in silico studies provide various ecological information. The genome is a reservoir for multiple heavy metals and metalloids resistance gene clusters as well as exhibit metabolic capabilities to utilize a wide range of carbohydrates. It also encodes for tris-catecholate siderophore and this can regulate uptake of iron and plant growth such as for mangrove vegetation.
How to Set up and Run a Bad Bugs Bookclub group: resources and recommendations
The Bad Bugs Bookclub is a public engagement initiative that enables scientists (microbiologists) and non-scientists to discuss the role of infectious disease and microorganisms in novels of fiction. The bookclub began in 2009 but since 2020 the meetings have taken place online enabling international membership and occasional author participation. The bookclub has been shown through peer-reviewed publications to have impact and value to its members. For each book (the number now exceeds 100) a reading guide (questions to provoke discussion) and a meeting report (narrative of the discussion) have been produced. Previously hosted on a website the reading guides from this rich archive and resource are now presented alongside this paper which provides tips on how to run a similar reading group.
Rising Clindamycin Resistance in Group A Streptococcus in an Irish Healthcare Institution
Group A streptococcus (GAS) can cause serious invasive disease in humans with a high mortality rate. An increase in GAS infections was reported in Ireland in 2022 and this increase has been sustained in 2023 and is paralleled by similar trends in Europe. Rising antimicrobial resistance is a global problem and presents significant challenges to clinicians treating GAS infection. There was a reported increase in clindamycin resistance in GAS isolates in Ireland in 2022. We examined antimicrobial susceptibility patterns of GAS isolates in our institution in 2022. Although all GAS isolates included in our study were susceptible to penicillin we noted a high clindamycin resistance rate of 28% in our invasive GAS isolates. We also noted high tetracycline and erythromycin resistance 43% and 30% respectively. Our results could have implications for empiric antimicrobial prescribing guidelines for skin and soft tissue infections which often include clindamycin as it inhibits the production of many virulence factors associated with GAS. In addition macrolides are often the first line recommended antibiotic for patients with anaphylaxis to penicillin. This study emphasises the importance of continuous surveillance and antimicrobial susceptibility testing of invasive and non-invasive isolates in order to monitor trends in increasing antimicrobial resistance.
Evaluation of Susceptibility Status of Some Clinical Bacterial Isolates to Generations of Cephalosporins Sold in Kano
Introduction: The circulation of substandard antibiotics is believed to aggravate the evil effect of Antibiotic resistance (AR) globally. AR knows no boundaries and pose a serious threat to existing antibiotics
Gap statement: Sabon Gari market is chosen due to reports of substandard antibiotics. It's a major distribution centre for pharmaceuticals yet research on its impact and extent of substandard drugs is lacking.
Aim: This study aims to evaluate the susceptibility status of some clinical Bacterial isolates (E. coli K. pneumoniae S. aureus S. pneumoniae P. aeruginosa and Salmonella sp.) to generations of Cephalosporins purchased from drug distributors in Sabon Gari Market Kano.
Methodology: Antimicrobial susceptibility pattern of the test isolates was determined by disc diffusion method. Fourier Transform Infrared Spectroscopy (FT-IR) analysis was used to confirm the functional group of the active ingredients of all the antibiotics tested. Molecular identification of resistant gene (CTX-M1) were carried out using PCR.
Results: Market survey reveals that Cephalexin 61% (first generation) Cefuroxime 72% (Second generation) and Cefexime 68% Cefpodoxime 79% Ceftriazone 63% Ceftazidime 70% and Cefotaxime 45% (third generation) and Cefepime 84% (fourth generation) were the most commonly sold Cephalosporins with different brands and company names. In all the antibiotics tested similar activity was observed in the branded antibiotics as compared with the standard antibiotics with no significant difference observed. Exactly 20% of E. coli and K. pneumoniae were resistant while 80% of S. aureus S. pneumoniae Pseudomonas and Salmonella sp. were susceptible. CTX-M1 resistant gene was identified E. coli and K. pneumoniae which further confirms their resistance to Cefotaxime and ceftriaxone antibiotics.
Conclusion: Branded cephalosporins sold in Kano were effective as standard antibiotics with good bioavailability and compliance with the standards formulation of medicine thus fit for human intake.
Identification of causative fungus from sterile abscess using metagenomics followed by in situ hybridization
Introduction: Invasive fungal infections require early diagnosis for treatment. Microscopic observation of biopsy and blood culture are the gold standard for identification of the causative fungus but it is difficult to identify the causative pathogen by a sterile abscess biopsy.
Case Presentation: We present a case report of breakthrough invasive trichosporonosis in a 65-year-old Japanese male with acute myeloid leukemia (AML) receiving antifungal prophylaxis. Blood cultures showed no fungal growth and a liver biopsy and a removed spleen with abscess showed fragmented fungi but no fungal identification was possible. This report demonstrates that retrospective analyses were able to identify the causative fungus.
Conclusion: We narrowed down the candidate fungi by deep sequencing of the ITS1 region of fungal genome and confirmed that the fungus observed in the specimen was T. asahii by in situ hybridization using a DNA probe targeting 26S rRNA.
Methods to assess antibacterial, antifungal and antiviral surfaces in relation to touch and droplet transfer: a review, gap-analysis and suggested approaches.
To help assess whether a potentially antimicrobial material surface or coating provides antimicrobial efficacy a number of standardised test methods have been developed internationally. Ideally these methods should generate data that supports the materials efficacy when deployed in the intended end-use application. These methods can be categorised based on their methodological approach such as suspension tests agar plate/zone diffusion tests surface inoculation tests surface growth tests or surface adhesion tests. To support those interested in antimicrobial coating efficacy this review brings together an exhaustive list of methods (for porous and non-porous materials) exploring the methodological and environmental parameters used to quantify antibacterial antifungal or viral activity. This analysis demonstrates that antimicrobial efficacy methods that test either fungi or viruses are generally lacking whilst methods that test bacteria fungi and viruses are not designed to simulate end-use/lack realistic conditions. As such a number of applications for antimicrobial activity across medical touch screens medical textiles and gloves and transport seat textiles are explored as example applications providing guidance on modifications to existing methods that may better simulate the intended end-use of antimicrobial materials.
Primary iliac bone tuberculosis: A case report
Tuberculosis is an infectious disease caused by the Mycobacterium tuberculosis complex. It is a major public health problem and one of the world's leading causes of morbidity and mortality. It occurs in both pulmonary and extra-pulmonary forms the pulmonary form being the most common. Primary iliac bone tuberculosis remains a rare clinical entity even in endemic areas. Its diagnosis can be challenging due to its similarity to other bone conditions. We report a rare case of primary iliac bone tuberculosis in a 63-year-old patient on peritoneal dialysis with the following medical history: hypertension type II diabetes complicated by diabetic retinopathy and diabetic kidney disease. Recent advances in molecular biology in particular with the advent of the Genexpert® have considerably improved patient management providing microbiological evidence in less than two hours.
Atypical presentation of varicella-zoster virus reactivation in a lung transplant patient: a case report
Background: Varicella zoster virus (VZV) is a human neurotropic virus which commonly causes infection during childhood. Later in life it may reactivate as herpes zoster. We report a rare manifestation of reactivation of VZV infection presenting as cutaneous vasculitis and varicella pneumonia in a lung transplant recipient.
Case presentation: A 65 year old man was lung transplanted bilaterally for emphysema and had repeated posttransplant chest infections and colonization with Pseudomonas aeruginosa. Nine months post-transplant he presented with dyspnea and a cutaneous vasculitis-like eruption. VZV reactivation was not suspected due to absence of the typical vesicular eruptions. The diagnosis was confirmed by VZV PCR from the swabs of the ulcer after skin punch biopsy of a lesion and from bronchoalveolar lavage (BAL). The histology of skin biopsy demonstrated epithelial and vascular damage but no typical epithelial virus associated changes. The patient responded to antiviral therapy with total remission of rash and VZV DNA was not detectable from repeated BAL after 29 days of therapy. However the pulmonary radiological features and dyspnea persisted due to reasons possibly unrelated to the VZV infection.
Conclusion: Had it not been for the patient to mention the resemblance of the vasculitic rash with his primary VZV infection the diagnosis would easily have been overlooked. In this case the biopsy did not show typical histopathologic findings of VZV-vasculitis. What led the diagnosis was PCR from the wound swab taken after punch biopsy. This case serves as a reminder for atypical presentation of common conditions in immunosuppressed patients and that extensive diagnostic sampling may be warranted in this group.
Sarcina Ventriculi in Association with Gastric Ulcer: A Case Report
Sarcina ventriculi is a gram positive bacteria which has been reported in patients with delayed gastric emptying as well as in association with cases of gastric ulcer and gastric carcinoma. Although it has been reported frequently in veterinary cases as a cause of fatal diseases exact pathogenesis in humans is yet to be identified. We report here a case of an elderly male who presented with hematemesis following which an upper gastrointestinal endoscopy was done and a gastric ulcer was revealed. Histopathological examination revealed Sarcina ventriculi in association with ulcer.
A clinical metagenomic study of biopsies from Mexican endophthalmitis patients reveals the presence of complex bacterial communities and a diversity of resistance genes
Infectious endophthalmitis is a severe ophthalmic emergency. It is known that this infection can be caused by bacteria and fungi. For efficient treatment the administration of antimicrobial drugs to which the microbes are susceptible is essential. The aim of this study was to identify microorganisms in biopsies of Mexican endophthalmitis patients using metagenomic next-generation sequencing and determine which antibiotic resistance genes were present in the biopsy samples. In this prospective case study 19 endophthalmitis patients were recruited. Samples of vitreous or aqueous humor were extracted for DNA extraction for metagenomic next-generation sequencing. Analysis of the sequencing results revealed the presence of a wide variety of bacteria in the biopsies. The resistome analysis showed that homologs of antibiotic resistance genes were present in several biopsy samples. Genes possibly conferring resistance to ceftazidime and vancomycin were detected in addition to various genes encoding efflux pumps. Our findings contrast with the widespread opinion that only one or a few bacterial strains are present in the infected tissues of endophthalmitis patients. These diverse communities might host many of the resistance genes that were detected which can further complicate the infections.
Genomic diversity of novel strains of mammalian gut microbiome derived Clostridium XIVa strains is driven by mobile genetic element acquisition
Despite advances in sequencing technologies that enable a greater understanding of mammalian gut microbiome composition our ability to determine a role for individual strains is hampered by our inability to isolate culture and study such microbes. Here we describe highly unusual Clostridium XIVa group strains isolated from the murine gut. Genome sequencing indicates that these strains Clostridium symbiosum LM19B and LM19R and Clostridium clostridioforme LM41 and LM42 have significantly larger genomes than most closely related strains. Genomic evidence indicates that the isolated LM41 and LM42 strains diverge from most other Clostridium XIVa strains and supports reassignment of these groups at genus-level. We attribute increased C. clostridioforme LM41 and LM42 genome size to acquisition of mobile genetic elements including dozens of prophages integrative elements putative group II introns and numerous transposons including 29 identical copies of the IS66 transposase and a very large 192 Kb plasmid. antiSmash analysis determines a greater number of biosynthetic gene clusters within LM41 and LM42 than in related strains encoding a diverse array of potential novel antimicrobial compounds. Together these strains highlight the potential untapped microbial diversity that remains to be discovered within the gut microbiome and indicate that despite our ability to get a top down view of microbial diversity we remain significantly blinded to microbe capabilities at the strain level.
Spectrum of Respiratory Viruses Identified from SARS-CoV-2 Negative Specimens in Watansoppeng, a Bat City in Eastern Indonesia
Respiratory infections account for millions of hospital admissions worldwide. Understanding the etiology could aid management and preventive strategies to reduce morbidity and mortality. Bats are reported as one of the animal reservoirs for many emerging respiratory viruses. SARS-CoV-2 negative specimens from Wattansoppeng city South Sulawesi a natural habitat for Acerodon celebensis and Pteropus alecto fruit bats were analyzed to study the spectrum of respiratory viruses. Samples were screened for influenza virus Enterovirus Paramyxoviridae Nipah virus Coronaviridae and Pneumoviridae. Of 210 specimens 19 were positive for Respiratory Syncytial Virus (RSV)-A RSV-B human parainfluenza (HPIV)-1 virus HPIV-2 human rhinovirus (HRV)-A HRV-B HRV-C human metapneumovirus (HMPV) influenza A virus and CV-A6. Influenza virus was of seasonal H3N2 subtype. The HMPVs were of genotype B1 and A2a while one of the RSV-A was ON-1 genotype. The viruses mostly affected children with unknown severity. No novel viruses were observed in this study.
Aspergillus esophagitis in a patient with solid tumors: a case report
Esophageal aspergillosis is a rare occurrence primarily documented in hematologic malignancies and only rarely occurring among patients with solid tumors. In this case report we present the unique case of an 81-year-old Lebanese man who had a remarkable medical history including four solid tumors. The patient sought medical attention due to dysphagia and weight loss prompting a gastroscopic examination that revealed a necrotic abscess at the esophagogastric junction. Initial treatment with fluconazole and Proton Pump Inhibitors was administered but the recurrence of similar symptoms led to a repeat gastroscopy unveiling a diagnosis of Aspergillus esophagitis. Intravenous Voriconazole was promptly initiated; however the patient developed a significant pericardial effusion and expired with Aspergillus species identified in the pericardial fluid. This exceptional case emphasizes the importance of considering esophageal aspergillosis in cancer patients who present with refractory symptoms such as epigastric pain dysphagia nausea and vomiting despite symptomatic treatment. Our findings underscore the need for increased awareness and the inclusion of gastrointestinal endoscopy as part of the diagnostic approach for this rare but potentially life-threatening condition.
Sequence and origin of the Streptomyces intergenetic-conjugation helper plasmid pUZ8002.
Conjugation of plasmids from Escherichia coli is essential for the genetic manipulation of Streptomyces spp. To facilitate intergeneric conjugation from E. coli to Streptomyces the conjugative machinery required for genetic transfer is usually provided by the non-transferable helper plasmid pUZ8002. Here we present the complete nucleotide sequence of pUZ8002 describe the previously undocumented creation process and provide details of the sequence relative to the parental pUZ8 plasmid and another previously published pUZ8002 sequence.
Enhancement of growth media for extreme iron-limitation in Escherichia coli
Iron is an essential nutrient for microbial growth and bacteria have evolved numerous routes to solubilise and scavenge this biometal which is often present at very low concentrations in host tissue. We recently used a MOPS-based medium to induce iron limitation in Escherichia coli K-12 during the characterisation of novel siderophore conjugated antibiotics. In this study we confirm that growth media derived from commercially-available M9 salts are unsuitable for studies of iron-limited growth likely through the contamination of the sodium phosphate buffer components with over 100 µM iron. In contrast MOPS-based media that are treated with metal-binding Chelex® resin allow the free iron concentration to be reduced to growth-limiting levels. Despite these measures a small amount of E. coli growth is still observed in these iron-depleted media. By growing E. coli in conditions that theoretically increase the demand for iron-dependent enzymes namely by replacing the glucose carbon source for acetate and by switching to a microaerobic atmosphere we can reduce background growth even further. Finally we demonstrate that by adding an exogeneous siderophore to the growth media which is poorly used by E. coli we can completely prevent growth perhaps mimicking situation in host tissue. In conclusion this short study provides practical experimental insight into low iron media and how to augment the growth conditions of E. coli for extreme iron-limited growth.
Characterization of Group A streptococci causing invasive diseases in Sri Lanka
Group A β haemolytic streptococci (GAS) or Streptococcus pyogenes is a human pathogen that causes an array of infections including pharyngitis cellulitis impetigo scarlet fever toxic shock syndrome and necrotizing fasciitis. The present study characterizes 51 GAS isolates from invasive infections in Sri Lanka focusing on resistance profiles genetic determinants of resistance and virulence markers.
Isolates were tested for sensitivity to penicillin erythromycin clindamycin and tetracycline. The presence of erm(A) erm(B) mef(A) was detected in erythromycin-resistant isolates while tet(M) was detected in the tetracycline-resistant isolates. PCR was used to identify SpeA SpeB SpeC SpeF SpeG smez and ssa as virulence markers. Selected GAS isolates were emm-typed using the updated CDC protocol.
All 51 isolates were susceptible to penicillin. The number of isolates non-susceptible to erythromycin was 16. The commonest resistant determinant identified was erm(B) (11/16). Tetracycline nonsusceptibility was found in 36 (70.6%) isolates and 26 of them contained the tet(M) gene. Thirteen (25.5%) isolates were resistant to both tetracycline and erythromycin while 12 (23.5%) isolates were sensitive to both antibiotics. The commonest virulence markers detected among the isolates was SpeB (44 86.3%) SpeG (36 70.6%) SpeF (35 68.6%) while SpeJ (15 29.4%) SpeA (10 19.6%) and ssa (59.8%) were less common.
In conclusion GAS isolates studied showed resistance to erythromycin and tetracycline while retaining universal susceptibility to penicillin. Additionally these isolates exhibited diverse genetic backgrounds displaying varying patterns of virulence genes and emm types.
A study on viruses and bacteria with particular interest on Mycoplasma pneumoniae in children with exacerbation of asthma from a tertiary care hospital in Sri Lanka
Asthma is a significant public health concern particularly in children with severe symptoms. Exacerbation of asthma (EOA) is life-threatening and respiratory infections (RIs) play a crucial role. Though viruses play a significant role in EOA patients are empirically treated with antibiotics which contribute to the development of antibiotic resistance. Although there are widely reported association of EOA with viral or M. pneumoniae infections there are no published data in Sri Lanka. The present study aimed to identify the association of common respiratory viruses typical respiratory bacterial pathogens and M. pneumoniae in children with EOA and relate them with the compatibility of antimicrobial use.
A case-control study was conducted in the pediatric unit of North Colombo Teaching Hospital Sri Lanka involving two groups of children between 5-15 years of age. Group-1: children with EOA Group-2: children with stable asthma (SA). Each group consisted of 100 children. Sputum/throat swabs were tested for common respiratory viruses using virus specific FITC-labelled monoclonal antibodies (MAbs) bacteria by routine culture and M. pneumoniae by RT-PCR. Macrolide-resistance in M. pneumoniae was detected using conventional PCR and sequencing specific genetic mutations in the 23S rRNA gene. M. pneumoniae was genotyped using nested multilocus sequence typing (MLST) which targeted eight housekeeping genes (ppa pgm gyrB gmk glyA atpA arcC adk).
There was no significant difference in age gender demographic or geographical locations between the two groups. In children with EOA antibiotics were used in 66% (66/100) and macrolides in 42% (42/100) in children with EOA. Samples consisted of 78% (78/100) sputum and 22% (22/100) throat swabs. Adenovirus was the most common virus identified and it was significantly higher in children with EOA compared to those with SA but no significant difference in typical bacteria findings between the two groups. M. pneumoniae was detected in one patient with EOA with none detected in the SA group. The M. pneumoniae was macrolide sensitive and it was ST14 by Multi Locus Sequence Typing. This study showed that the empiric use of antibiotics in children with asthma may be better targeted with prior pathogen screening to inform appropriate treatment to minimize antibiotic resistance.
The Y498T499-SARS-CoV-2 Spike (S) protein variant interacts with rat ACE2 but does not infect or induce responses in the rat lung when delivered as a S-protein pseudotyped lentivirus
The rat is a useful laboratory model for respiratory disease and SARS-CoV-2 proteins such as the spike (S) protein can induce inflammation. This study has investigated the ability of the Q498Y P499T (QP-YT) amino acid change described in the S-protein of the mouse adapted laboratory SARS-CoV-2 MA strain to interact with rat angiotensin converting enzyme-2 (ACE2) and stimulate responses in rat lung. Using a real-time S-ACE2 quantitative fusion assay ancestral S-protein fuses with human but not rat ACE2. The QP-YT S-protein retains ability to fuse with human ACE2 and interacts with rat ACE2 in the fusion assay and using a S-protein pseudotyped lentivirus infection system. L452R S-protein did not bind to rat ACE2. Although rat lower lung contains both ACE2 and TMPRSS2 target cells intratracheal delivery of ancestral or QP-YT S-protein pseudotyped lentivirus did not induce measurable respiratory changes inflammatory infiltration or innate mRNA responses. Isolation of primary cells from rat alveoli demonstrated the presence of cells expressing ACE2 and TMPRSS2. Infection of these cells however with ancestral or QP-YT S-protein pseudotyped lentivirus was not observed. Analysis of the amino acid changes across the S-ACE2 interface highlights the Y498 interaction with H353 as a likely facilitator of binding to rat ACE2 but also other amino acids that could improve this interaction.
Thus rat lungs contain cells expressing receptors for SARS-CoV-2 and the QP-YT S-protein variant can bind to rat ACE2 but this does not result in infection or stimulate responses in the lung. Further amino acid changes in S-protein could enhance this interaction to improve the utility of the rat model for defining the role of the S-protein in driving inflammation in the lung.