The African trypanosome Trypanosoma brucei monoallelically expresses one of more than 1000 Variant Surface Glycoprotein (VSG) genes. The active VSG is transcribed from one of about 15 telomeric VSG expression sites (ESs). It is unclear how monoallelic expression of VSG is controlled, and how inactive VSG ESs are silenced. Here, we show that blocking synthesis of the T. brucei FACT subunit TbSpt16 triggers a G2/early M phase cell cycle arrest in both bloodstream and insect form T. brucei. Segregation of T. brucei minichromosomes in these stalled cells is impaired, implicating FACT in maintenance of centromeres. Strikingly, knock-down of TbSpt16 results in 20- to 23-fold derepression of silent VSG ES promoters in bloodstream form T. brucei, with derepression specific to the G2/M cell cycle stage. In insect form T. brucei TbSpt16 knock-down results in 16- to 25-fold VSG ES derepression. Using chromatin immunoprecipitation (ChIP), TbSpt16 was found to be particularly enriched at the promoter region of silent but not active VSG ESs in bloodstream form T. brucei. The chromatin remodeler FACT is therefore implicated in maintenance of repressed chromatin present at silent VSG ES promoters, but is also essential for chromosome segregation presumably through maintenance of functional centromeres.

Trypanosoma brucei mono-allelically expresses one of approximately 1500 variant surface glycoprotein (VSG) genes while multiplying in the mammalian bloodstream. The active VSG is transcribed by RNA polymerase I in one of approximately 15 telomeric VSG expression sites (ESs). T. brucei is unusual in controlling gene expression predominantly post-transcriptionally, and how ESs are mono-allelically controlled remains a mystery. Here we identify a novel transcription regulator, which resembles a nucleoplasmin-like protein (NLP) with an AT-hook motif. NLP is key for ES control in bloodstream form T. brucei, as NLP knockdown results in 45- to 65-fold derepression of the silent VSG221 ES. NLP is also involved in repression of transcription in the inactive VSG Basic Copy arrays, minichromosomes and procyclin loci. NLP is shown to be enriched on the 177- and 50-bp simple sequence repeats, the non-transcribed regions around rDNA and procyclin, and both active and silent ESs. Blocking NLP synthesis leads to downregulation of the active ES, indicating that NLP plays a role in regulating appropriate levels of transcription of ESs in both their active and silent state. Discovery of the unusual transcription regulator NLP provides new insight into the factors that are critical for ES control.

The assistantship seems to have been highly valued by students, but could be improved by ensuring that all students are given relevant placements and clinical responsibility.

Background This study aimed to decrease unnecessary long-term antimicrobial use in primary care by identifying and highlighting patients with long-term or repeated exposure to antibiotics to GP teams for review. The broad spectrum antimicrobial co-amoxiclav was targeted following data showing rising DDD per prescription in our Health Board. Methods Community antibiotic prescribing from the national Prescribing Information System (PIS) was searched for NHS Lothian (126 GP practices) using DDD to identify patients. Long courses of co-amoxiclav were defined as ≥63 DDD. Ninety-eight patients were identified with high antimicrobial usage and a data collection form was sent to the Primary Care Pharmacy teams to voluntarily collate data for each patient. Results The data collection form had a 97% response rate. Of the 94 patients with full data 62 (66%) patients had co-amoxiclav initiated due to secondary care specialist advice and 26 (28%) had been initiated by primary care. Forty-four patients (47%) were aged 65 or over, putting them at high risk of Clostridioides difficile. Forty-one patients had multiple courses of co-amoxiclav and 39 patients were prescribed co-amoxiclav as prophylaxis; of these 59% were for urinary tract infection (UTI). Thirty-four of 94 patients (36%) had been on prophylaxis for over 5 years with 9 patients on co-amoxiclav for over 20 years. Of note in 39 of 94 patients no microbiology samples were available and where samples were available (n=55), narrower spectrum options were available in 38 patients (69%) and 17 patients (31%) had samples showing resistance to co-amoxiclav. Following this intervention to highlight use, the number of patients on long-term courses or prophylaxis with co-amoxiclav reduced by 70% in the following year. Conclusions Patients on long-term antibiotics are priorities for antimicrobial stewardship interventions. This work demonstrated that ongoing appropriateness and efficacy of long-term antibiotic courses were not routinely reviewed. Using pharmacy data proved an effective method to highlight patients and ultimately reduce patient antibiotic exposure. UTI prophylaxis was the majority patient group in this study and interventions to review and decrease long-term UTI prophylaxis in our Board using this method are underway.

In the article by Denninger et al. (2010), the affiliation of the third co-author Mohamed Bessat has been updated to include a missing affiliation. ReferenceDenninger, V., Fullbrook, A., Bessat, M., Ersfeld, K., and Rudenko, G. (2010) The FACT subunit TbSpt16 is involved in cell cycle specific control of VSG expression sites in Trypanosoma brucei. Molecular Microbiology 78 (2): 459-474.