The TIR-domain containing effectors BtpA and BtpB from Brucella abortus impact NAD metabolism.

Brucella species are facultative intracellular Gram-negative bacteria relevant to animal and human health. Their ability to establish an intracellular niche and subvert host cell pathways to their advantage depends on the delivery of bacterial effector proteins through a type IV secretion system. Brucella Toll/Interleukin-1 Receptor (TIR)-domain-containing proteins BtpA (also known as TcpB) and BtpB are among such effectors. Although divergent in primary sequence, they interfere with Toll-like receptor (TLR) signaling to inhibit the innate immune responses. However, the molecular mechanisms implicated still remain unclear. To gain insight into the functions of BtpA and BtpB, we expressed them in the budding yeast Saccharomyces cerevisiae as a eukaryotic cell model. We found that both effectors were cytotoxic and that their respective TIR domains were necessary and sufficient for yeast growth inhibition. Growth arrest was concomitant with actin depolymerization, endocytic block and a general decrease in kinase activity in the cell, suggesting a failure in energetic metabolism. Indeed, levels of ATP and NAD+ were low in yeast cells expressing BtpA and BtpB TIR domains, consistent with the recently described enzymatic activity of some TIR domains as NAD+ hydrolases. In human epithelial cells, both BtpA and BtpB expression reduced intracellular total NAD levels. In infected cells, both BtpA and BtpB contributed to reduction of total NAD, indicating that their NAD+ hydrolase functions are active intracellularly during infection. Overall, combining the yeast model together with mammalian cells and infection studies our results show that BtpA and BtpB modulate energy metabolism in host cells through NAD+ hydrolysis, assigning a novel role for these TIR domain-containing effectors in Brucella pathogenesis.

Studying Coxiella burnetii Type IV Substrates in the Yeast Saccharomyces cerevisiae: Focus on Subcellular Localization and Protein Aggregation.

Coxiella burnetii is a Gram-negative obligate parasitic bacterium that causes the disease Q-fever in humans. To establish its intracellular niche, it utilizes the Icm/Dot type IVB secretion system (T4BSS) to inject protein effectors into the host cell cytoplasm. The host targets of most cognate and candidate T4BSS-translocated effectors remain obscure. We used the yeast Saccharomyces cerevisiae as a model to express and study six C. burnetii effectors, namely AnkA, AnkB, AnkF, CBU0077, CaeA and CaeB, in search for clues about their role in C. burnetii virulence. When ectopically expressed in HeLa cells, these effectors displayed distinct subcellular localizations. Accordingly, GFP fusions of these proteins produced in yeast also decorated distinct compartments, and most of them altered cell growth. CaeA was ubiquitinated both in yeast and mammalian cells and, in S. cerevisiae, accumulated at juxtanuclear quality-control compartments (JUNQs) and insoluble protein deposits (IPODs), characteristic of aggregative or misfolded proteins. AnkA, which was not ubiquitinated, accumulated exclusively at the IPOD. CaeA, but not AnkA or the other effectors, caused oxidative damage in yeast. We discuss that CaeA and AnkA behavior in yeast may rather reflect misfolding than recognition of conserved targets in the heterologous system. In contrast, CBU0077 accumulated at vacuolar membranes and abnormal ER extensions, suggesting that it interferes with vesicular traffic, whereas AnkB associated with the yeast nucleolus. Both effectors shared common localization features in HeLa and yeast cells. Our results support the idea that C. burnetii T4BSS effectors manipulate multiple host cell targets, which can be conserved in higher and lower eukaryotic cells. However, the behavior of CaeA and AnkA prompt us to conclude that heterologous protein aggregation and proteostatic stress can be a limitation to be considered when using the yeast model to assess the function of bacterial effectors.

Patrones de resistencia de Neisseria gonorrhoeae aisladas en el área de Cuenca / Resistance patterns of Neisseria gonorrhoeae isolated in patients from the area of cuenca (Spain)

El artículo es un estudio retrospectivo para conocer la sensibilidad antibiótica de los aislados de Neisseria gonorhoeae de muestras genitales de pacientes durante los últimos 4 años en el área de Cuenca (España) para así establecer cuáles serían las mejores pautas de tratamiento para esta enfermedad en nuestra región. De un total de 26 cepas aisladas de Neisseria gonorrhoeae, la resistencia a fluorquinolonas fue del 61,54%. Todas las cepas fueron sensibles a cefotaxima, por lo que consideramos que el tratamiento empírico de elección de las infecciones gonocócicas no complicadas en nuestro medio debería ser las cefalosporinas de tercera generación (AU)

A retrospective study was conducted to determine the antibiotic susceptibility of Neisseria gonorrhoeae isolates from genital samples of patients over the last four years in Cuenca (Spain), in order to establish what would be the best treatment regimens for this disease in our region. Of a total of 26 isolates of Neisseria gonorrhoeae, fluoroquinolone resistance was 61.54%. All strains were susceptible to cefotaxime, so we believe that the choice of empirical treatment of uncomplicated gonococcal infections should be third generation cephalosporins in our region (AU)

Prevalencia de parasitosis intestinales en niños de acogida saharauis / Prevalence of intestinal parasite infestation in foster children from the Sahara

El artículo es un estudio retrospectivo para conocer la prevalencia de parasitaciones intestinales en niños de acogida saharauis en la provincia de Cuenca (España). Se incluyeron un total de 157 muestras de 90 pacientes durante un período de cinco años. Encontramos que los pacientes presentaban un porcentaje de parasitación del 37,37%. Sería conveniente realizar un protocolo para el diagnóstico y tratamiento de las parasitaciones intestinales en estos niños (AU)

We performed a retrospective study to find out the prevalence of intestinal parasite infestation in foster children from the Sahara in Cuenca (Spain). We included 157 samples from 90 patients over a five-year period. It was found that 37.37% of the children were infected with pathological parasites. It would be advisable to develop a protocol for the diagnosis and treatment of intestinal parasite infestation in these children (AU)