Optically Mapping Multiple Bacterial Genomes Simultaneously in a Single Run

Optical mapping of bacterial chromosomes provides an unambiguous low-resolution sequence scaffold of the entire chromosome. In comparison to some techniques, such as pulse field gel electrophoresis, cost and throughput limit the application of this technique outside of genome finishing. We have demonstrated the production of multiple bacterial maps using a single set of consumables; this significantly reduces the time and expense of map production

Bacterial Peritonitis Due to \u3ci\u3eAcinetobacter baumannii\u3c/i\u3e Sequence Type 25 with Plasmid-Borne New Delhi Metallo-Beta-Lactamase in Honduras

A carbapenem-resistant Acinetobacter baumannii strain was isolated from the peritoneal fluid of a patient with complicated intra-abdominal infection and evaluated at the Multidrug-resistant Organism Repository and Surveillance Network by wholegenome sequencing and real-time PCR. The isolate was sequence type 25 and susceptible to colistin and minocycline, with low MICs of tigecycline. blaNDM-1 was located on a plasmid with \u3e99% homology to pNDM-BJ02. The isolate carried numerous other antibiotic resistance genes, including the 16S methylase gene, armA

Detection of New Delhi Metallo-β-Lactamase (Encoded by \u3ci\u3ebla\u3c/i\u3e\u3csub\u3eNDM-1\u3c/sub\u3e) in \u3ci\u3eAcinetobacter schindleri\u3c/i\u3e during Routine Surveillance

A carbapenem-resistant Alcaligenes faecalis strain was isolated from a surveillance swab of a service member injured in Afghanistan. The isolate was positive for blaNDM by real-time PCR. Species identification was reevaluated on three identification systems but was inconclusive. Genome sequencing indicated that the closest relative was Acinetobacter schindleri and that blaNDM-1 was carried on a plasmid that shared \u3e99% identity with one identified in an Acinetobacter lwoffii isolate. The isolate also carried a novel chromosomally encoded class D oxacillinase

Rapid whole genome optical mapping of Plasmodium falciparum

<p>Abstract</p> <p>Background</p> <p>Immune evasion and drug resistance in malaria have been linked to chromosomal recombination and gene copy number variation (CNV). These events are ideally studied using comparative genomic analyses; however in malaria these analyses are not as common or thorough as in other infectious diseases, partly due to the difficulty in sequencing and assembling complete genome drafts. Recently, whole genome optical mapping has gained wide use in support of genomic sequence assembly and comparison. Here, a rapid technique for producing whole genome optical maps of <it>Plasmodium falciparum </it>is described and the results of mapping four genomes are presented.</p> <p>Methods</p> <p>Four laboratory strains of <it>P. falciparum </it>were analysed using the Argus™ optical mapping system to produce ordered restriction fragment maps of all 14 chromosomes in each genome. <it>Plasmodium falciparum </it>DNA was isolated directly from blood culture, visualized using the Argus™ system and assembled in a manner analogous to next generation sequence assembly into maps (AssemblyViewer™, OpGen Inc.^®). Full coverage maps were generated for <it>P. falciparum </it>strains 3D7, FVO, D6 and C235. A reference <it>P. falciparum in silico </it>map was created by the digestion of the genomic sequence of <it>P. falciparum </it>with the restriction enzyme AflII, for comparisons to genomic optical maps. Maps were then compared using the MapSolver™ software.</p> <p>Results</p> <p>Genomic variation was observed among the mapped strains, as well as between the map of the reference strain and the map derived from the putative sequence of that same strain. Duplications, deletions, insertions, inversions and misassemblies of sizes ranging from 3,500 base pairs up to 78,000 base pairs were observed. Many genomic events occurred in areas of known repetitive sequence or high copy number genes, including <it>var </it>gene clusters and <it>rifin </it>complexes.</p> <p>Conclusions</p> <p>This technique for optical mapping of multiple malaria genomes allows for whole genome comparison of multiple strains and can assist in identifying genetic variation and sequence contig assembly. New protocols and technology allowed us to produce high quality contigs spanning four <it>P. falciparum </it>genomes in six weeks for less than $1,000.00 per genome. This relatively low cost and quick turnaround makes the technique valuable compared to other genomic sequencing technologies for studying genetic variation in malaria.</p

A Double-Blind Randomized Phase I Clinical Trial Targeting ALVAC-HIV Vaccine to Human Dendritic Cells

BACKGROUND: We conducted a novel pilot study comparing different delivery routes of ALVAC-HIV (vCP205), a canarypox vaccine containing HIV gene inserts: env, gag and pol. We explored the concept that direct ex vivo targeting of human dendritic cells (DC) would enhance the immune response compared to either conventional intramuscular or intradermal injections of the vaccine alone. METHODOLOGY/PRINCIPAL FINDINGS: Healthy HIV-1 uninfected volunteers were administered ALVAC-HIV or placebo by intramuscular injection (i.m.), intradermal injection (i.d.) or subcutaneous injection (s.q.) of autologous ex vivo transfected DC at months 0, 1, 3 and 6. All vaccine delivery routes were well tolerated. Binding antibodies were observed to both the ALVAC vector and HIV-1 gp160 proteins. Modest cellular responses were observed in 2/7 individuals in the DC arm and 1/8 in the i.m. arm as determined by IFN-γ ELISPOT. Proliferative responses were most frequent in the DC arm where 4/7 individuals had measurable responses to multiple HIV-1 antigens. Loading DC after maturation resulted in lower gene expression, but overall better responses to both HIV-1 and control antigens, and were associated with better IL-2, TNF-α and IFN-γ production. CONCLUSIONS/SIGNIFICANCE: ALVAC-HIV delivered i.m., i.d. or s.q. with autologous ex vivo transfected DC proved to be safe. The DC arm was most immunogenic. Proliferative immune responses were readily detected with only modest cytotoxic CD8 T cell responses. Loading mature DC with the live viral vaccine induced stronger immune responses than loading immature DC, despite increased transgene expression with the latter approach. Volunteers who received the autologous vaccine loaded mature DC developed a broader and durable immune response compared to those vaccinated by conventional routes. TRIAL REGISTRATION: ClinicalTrials.gov NCT00013572

High Consequence Pathogens and HAI* (Healthcare Associated Infection) Priorities: Infectious Disease & Infectious Control Updates for 2022-2023

Rochester General Hospital Medical Grand Rounds for 2022-09-08, Dr. Emil Lesho, MD, Infectious Disease Division: Objectives: #1 Describe how to identify, isolate, confirm, and inform a monkey pox patient. #2 List the most important HAI* reduction goal for 2022/23 #3 Recognize emerging urgent public health threats such as candida avris and others in NYS & RRH

The Slow-Motion Catastrophe of Antimicrobial Resistance and Practical Interventions for All Prescribers

All medical and surgical specialties depend on the pool of effective antibiotics that continues to evaporate because of the increasing prevalence of drug-resistant bacteria. Antimicrobial-resistant infections kill 700,000 patients every year. By 2050, they are projected to cause 10 million deaths per year at a cumulative global cost of $100 trillion. Professional societies and international health agencies, including the United Nations, have declared escalating antimicrobial resistance as one of the gravest and most urgent threats to global public health and issued calls for action. The propensity of bacteria to mobilize and share genetic resistance determinants across species and genera, record levels of conflict-driven human population displacement, and the dearth of new antibiotics and rapid diagnostic tests, along with climate change and the epidemic of opioid addiction, exacerbate the antimicrobial resistance crisis. The predominant cause of antibiotic resistance is exposure to antibiotics through appropriate and inappropriate use. Mindfulness, nudging by peers, and adjuncts and alternatives to antibiotics, such as phage therapies, microbiome-based therapies, and novel medical informatics applications, could help reduce antibiotic use. This article describes the antimicrobial resistance crisis and highlights points in the continuum of care in which clinicians can readily implement practical, no-cost changes to minimize antibiotic exposure

The Rationale for a Mandatory Antibiotic Stewardship Rotation During Residency: What Worked, What Needs More Work

Reports of antibiotic stewardship (AS) integration into the \u3e 1000 United States internal medicine and family practice residency core curricula are scarce, but residents value such training. To help address this gap, and the projected shortage of physicians with training for establishing and leading an AS program (ASP), we describe the rationale for, and the output and shortcomings of, a dedicated AS rotation. Residents critically review, in real-time, inpatient antibiotic orders, provide feedback to the prescribers, learn the mechanics and requirements of an ASP, and complete a preliminary quality improvement project. Program evaluations are uniformly positive, noting limited opportunities otherwise to clarify optimal antibiotic choices or discuss antibiotics in depth. Nine posters at national conferences and 1 publication have roots in this rotation. Three alumni matriculated to accredited US infectious diseases fellowships. We invite others to join us in calling for more AS training opportunities during residency

Severe recurrent Streptobacillus moniliformis endocarditis in a pregnant woman, and review of the literature

BACKGROUND: Rat bite fever is a systemic febrile illness caused by infection with the Gram-negative bacillus Streptobacillus moniliformis following a bite, scratch, or contact with excrement. Only 26 cases of native valve endocarditis have been reported to date. We could find no other reports of severe Streptobacillus endocarditis requiring valve replacement in a young, pregnant patient. CASE PRESENTATION: A pregnant patient sought care for right leg pain, fevers, left upper quadrant pain, generalized weakness, fatigue, and inability to bear weight on her right leg. She had a syncopal episode 9 months earlier, resulting in a mandibular fracture and internal fixation hardware. Her pregnancy was complicated by hyperemesis and weight loss. Her pets included a rescued wild bird, a cat, and four rats. Her parents rescued stray cats, and she recalled multiple cat bites and scratches since childhood. She denied injection drug use. Ultrasound indicated a right popliteal artery thrombus. Transesophageal echocardiogram revealed a 2 cm × 0.7 cm vegetation. Angiography demonstrated multiple splenic infarcts and bilateral renal infarcts. She underwent mitral valve repair. The mitral valve Gram stain demonstrated 2+ Gram-negative rods, rare Gram-positive rods, and moderate white blood cells. Propionibacterium spp. was isolated from the mitral valve tissue on Columbia agar incubated anaerobically. Anaerobic and aerobic cultures of the valve tissue on all other broths and agars remained negative at 14 days. Hematoxylin and eosin stains showed a fibro-inflammatory vegetation. Aggregates of rod-shaped bacteria were identified on Warthin Starry/Steiner stain. Bartonella titers were positive for B. henselae IgG 1:256, IgM \u3c 1:20. Brown-Hopps Gram stain, AFB, and GMS stains for bacterial and fungal microorganisms were negative. Broad range bacterial PCR and sequencing of a segment of 16 s rRNA gene of the valve tissue matched to Streptobacillus sp. (genus level) and most closely related to Streptobacillus moniliformis. CONCLUSION: This case demonstrates diagnostic and therapeutic challenges associated with a relatively uncommon cause of endocarditis. The diagnosis of rat bite fever was delayed due to symptoms of a concomitant pregnancy. Other confounders included possible alternative sources or co-infections with another zoonosis from multiple pets, and an odontogenic source due to presence of exposed jaw hardware