scRNA-seq assessment of the human lung, spleen, and esophagus tissue stability after cold preservation.

BACKGROUND: The Human Cell Atlas is a large international collaborative effort to map all cell types of the human body. Single-cell RNA sequencing can generate high-quality data for the delivery of such an atlas. However, delays between fresh sample collection and processing may lead to poor data and difficulties in experimental design. RESULTS: This study assesses the effect of cold storage on fresh healthy spleen, esophagus, and lung from ≥ 5 donors over 72 h. We collect 240,000 high-quality single-cell transcriptomes with detailed cell type annotations and whole genome sequences of donors, enabling future eQTL studies. Our data provide a valuable resource for the study of these 3 organs and will allow cross-organ comparison of cell types. We see little effect of cold ischemic time on cell yield, total number of reads per cell, and other quality control metrics in any of the tissues within the first 24 h. However, we observe a decrease in the proportions of lung T cells at 72 h, higher percentage of mitochondrial reads, and increased contamination by background ambient RNA reads in the 72-h samples in the spleen, which is cell type specific. CONCLUSIONS: In conclusion, we present robust protocols for tissue preservation for up to 24 h prior to scRNA-seq analysis. This greatly facilitates the logistics of sample collection for Human Cell Atlas or clinical studies since it increases the time frames for sample processing.

The renal channelopathies.

Specific channels permit movement of selected ions through cellular membranes, and are of vital importance in a number of physiological processes, particularly in excitable tissues such as nerve and muscle, but also in endocrine organs and in epithelial biology. Disorders of channel proteins are termed channelopathies, and their importance is increasingly recognised within medicine. In the kidney, ion channels have critical roles enabling sodium and potassium reuptake or excretion along the nephron, in magnesium homeostasis, in the control of water reabsorption in the collecting duct, and in determining glomerular permeability. In this review, we assess the channelopathies encountered in each nephron segment, and see how their molecular and genetic characterisation in the past 20-30 years has furthered our understanding of normal kidney physiology and disease processes, aids correct diagnosis and promises future therapeutic opportunities.

Review of the clinical activity of medical microbiologists in a teaching hospital.

BACKGROUND: The clinical interactive role of medical microbiologists has been underestimated and the discipline is perceived as being confined to the laboratory. Previous studies have shown that most microbiology interaction takes place over the telephone. AIM: To determine the proportion of clinical ward based and laboratory based telephone interactions and specialties using a microbiology service. METHODS: Clinical microbiology activity that took place during November 1996 was prospectively analysed to determine the distribution of interactions and specialties using the service. RESULTS: In all, 1177 interactions were recorded, of which nearly one third (29%) took place at the bedside and 23% took place on call. Interactions involving the intensive treatment unit, general ward visits, and communication of positive blood cultures and antibiotic assays were the main areas of activity identified. There were 147 visits to 86 patients on the general wards during the study, with the number of visits to each individual varying from one to eight. The need for repeated visits reflected the severity of the underlying condition of the patients. Ward visits were regarded as essential to obtain missing clinical information, to assess response to treatment, and to make an appropriate entry in a patient's notes. CONCLUSIONS: Ward visits comprise a significant proportion of clinical microbiology interactions and have potential benefits for patient management, service utilisation, and education.

A case of peritonitis caused by Roseomonas gilardii in a patient undergoing continuous ambulatory peritoneal dialysis.

A case of peritonitis caused by Roseomonas gilardii in a patient receiving continuous ambulatory peritoneal dialysis is presented. The patient's domestic water supply was implicated as the probable source of infection. This is the first report of R. gilardii causing such an infection.

An epidemiological study of blood culture isolates of coagulase-negative staphylococci demonstrating hospital-acquired infection.

We applied pulsed-field gel electrophoresis (PFGE) after SmaI digestion and random amplification of polymorphic DNA (RAPD) analysis with nine oligonucleotide primers to 146 blood culture isolates of Staphylococcus epidermidis and 25 blood culture isolates of Staphylococcus haemolyticus. These were obtained over a 12-month period from patients on the neonatal and hematology units of the Central Manchester Health Care Trust. PFGE demonstrated two clusters of isolates of S. epidermidis (type A and type B) on the neonatal ward and a single cluster (type C) on the hematology unit. Type A was represented by 10 indistinguishable isolates from nine patients, type B was represented by 20 isolates from 14 patients, and type C was represented by 26 isolates from 10 patients. Type A isolates were resistant to chloramphenicol and type C isolates were resistant to ciprofloxacin, mirroring current antibiotic usage. There was no evidence of cross infection due to S. haemolyticus. RAPD analysis, on the basis of a single band difference, produced 58 types of S. epidermidis and 12 types of S. haemolyticus with primer 8 (ATG TAA GCT CCT GGG GAT TCA C; 5' to 3') and 54 types of S. epidermidis and 10 types of S. haemolyticus with primer 9 (AAG TAA GTG ACT GGG GTG AGC G; 5' to 3'). Combining the results confirmed cross infection. Types A, B, and C were concurrently isolated from the hands of the staff of the appropriate unit. Partial control was achieved by withdrawing ciprofloxacin use in the case of the hematology unit and improving hand hygiene in both units.

Kitchens as a source of Aspergillus niger infection.

In this study we investigated the epidemiology of a cluster of cutaneous infections owing to Aspergillus niger, which occurred in neutropenic patients in a bone marrow transplant unit. Heavy environmental contamination with the mould was found in the ward kitchen adjacent to the unit. The clinical and environmental isolates were typed by random amplification of polymorphic DNA (RAPD), which showed one of the patients was infected with the same strain as that isolated repeatedly from the kitchen area. In another case, contaminated stockinette material was implicated as the source of infection. Thorough cleaning of the ward kitchen resulted in no further cases on the unit. This highlights the fact that aspergilli may spread to patients by air, food or other vehicles, and underlines the importance of searching for a source and ensuring high levels of hospital hygiene are maintained.

"Pseudoclusters" and typing by random amplification of polymorphic DNA of Aspergillus fumigatus.

This short report serves as a warning to the unwary of possible "pseudoclusters" of infection with Aspergillus fumigatus as shown by the typing system, random amplification of polymorphic DNA (RAPD). This was demonstrated by typing 10 epidemiologically distinct isolates of A fumigatus using two different preparations of Taq DNA polymerase. One of the enzymes did not discriminate between the isolates, giving the false impression that a cluster of infection had occurred. Enzyme source is thus a key variable when using RAPD to distinguish between isolates of A fumigatus.

Invasive aspergillosis: clusters and sources?

Clusters of invasive infection with Aspergillus fumigatus are known to be associated with building works but studying the epidemiology has been hampered by the lack of a reliable typing system. A combination of three typing systems; silver staining of sodium dodecyl sulphate-polyacrylamide gels, immunoblot fingerprinting, and random amplification of polymorphic DNA (RAPD) was applied to seven cases on a haematology unit. The results show three of the patients to have indistinguishable isolates, suggesting a common source. Detection and removal of such sources, although difficult, would be an effective way of controlling the infection.

Application of polymerase chain reaction to fingerprinting Aspergillus fumigatus by random amplification of polymorphic DNA.

A new method for fingerprinting Aspergillus fumigatus by random amplification of polymorphic DNA (RAPD) by using single primers with arbitrary sequences is described. Five primers were examined with 19 isolates from six patients with aspergilloma as well as with A. fumigatus NCPF 2109. Two of the primers (GCT GGT GG and GCG CAC GG, 5' to 3') gave adequate discrimination between isolates, generating five and six types, respectively. Combination of the results obtained with each of these two primers generated 12 types. This compares very favorably with immunoblot fingerprinting and XbaI-generated restriction fragment length polymorphisms on the same isolates. Typeability and reproducibility were good with RAPD, and RAPD was less labor-intensive than immunoblot fingerprinting. RAPD typing results suggested that aspergillomas sometimes contain isolates of more than one type.