PD-L1 expression by dendritic cells is a key regulator of T-cell immunity in cancer.

Inhibiting the programmed death-1 (PD-1) pathway is one of the most effective approaches to cancer immunotherapy, but its mechanistic basis remains incompletely understood. Binding of PD-1 to its ligand PD-L1 suppresses T-cell function in part by inhibiting CD28 signaling. Tumor cells and infiltrating myeloid cells can express PD-L1, with myeloid cells being of particular interest as they also express B7-1, a ligand for CD28 and PD-L1. Here we demonstrate that dendritic cells (DCs) represent a critical source of PD-L1, despite being vastly outnumbered by PD-L1+ macrophages. Deletion of PD-L1 in DCs, but not macrophages, greatly restricted tumor growth and led to enhanced antitumor CD8+ T-cell responses. Our data identify a unique role for DCs in the PD-L1-PD-1 regulatory axis and have implications for understanding the therapeutic mechanism of checkpoint blockade, which has long been assumed to reflect the reversal of T-cell exhaustion induced by PD-L1+ tumor cells.

Detection of Diarrheagenic Escherichia coli in Bovine Meat in the Northern Region of Paraná State, Brazil

Abstract Ground bovine meat is commonly consumed by the population of Brazil. However, it constitutes an excellent medium for the multiplication of microorganisms due to available nutrients and handling practices prior to consumption. Here, we examined 100 samples of ground beef for the presence of diarrheagenic Escherichia coli (DEC) pathotypes by PCR, and characterized isolates by analyzing their adherence to HEp-2 cells, serotype, antimicrobial susceptibility, and phylogeny. Enteroaggregative E. coli was detected in five (5%) meat samples, Shiga toxin-producing E. coli in three (3%), and atypical enteropathogenic E. coli in two (2%). According to the phylogeny, six isolates (60%) were classified in group A, two (20%) in group B1, and two (20%) in group E. The detected serotypes were O3:H2, O93:H9, O93:H46, O105ab:H7, O152:H8, O156:H10, and O175:H7. The antimicrobial susceptibility testing showed that one sample (10%) was resistant to ampicillin, two (20%) to sulfamethoxazole-trimethoprim, and two (20%) to cephalothin. Based on these results, bovine ground meat for human consumption can serve as a reservoir of DEC, which emphasizes the importance of appropriate hygienic-sanitary conditions during handling at every stage from slaughter to table.

Diarrheagenic Escherichia coli Associated with Acute Gastroenteritis in Children from Soriano, Uruguay.

INTRODUCTION: Acute diarrheal disease still deserves worldwide attention due to its high morbidity and mortality, especially in developing countries. While etiologic determination is not mandatory for management of all individual cases, it is needed for generating useful epidemiologic knowledge. Diarrheagenic Escherichia coli (DEC) are relevant enteropathogens, and their investigation requires specific procedures to which resources and training should be dedicated in reference laboratories. METHODOLOGY: Following the hypothesis that enteric pathogens affecting children in towns located in the interior of Uruguay may be different from those found in Montevideo, we conducted a diagnostic survey on acute diarrheal disease in 83 children under 5 years of age from populations in the south of the country. RESULTS: DEC pathotypes were the only bacterial pathogens found in diarrheal feces (20.48%), followed by rotavirus (14.45%) and enteric adenovirus (4.81%). Atypical EPEC (aEPEC) was the most frequent DEC pathotype identified, and unexpectedly, it was associated with bloody diarrheal cases. These patients were of concern and provided with early consultation, as were children who presented with vomiting, which occurred most frequently in rotavirus infections. aEPEC serotypes were diverse and different from those previously reported in Montevideo children within the same age group and different from serotypes identified in regional and international studies. Enteroinvasive (EIEC) O96 : H19, associated with large outbreaks in Europe, was also isolated from two patients. Antibiotic susceptibility of pathogenic bacteria identified in this study was higher than that observed in previous national studies, which had been mainly carried out in children from Montevideo. CONCLUSION: The reduced number of detected species, the marked prevalence of aEPEC, the scarce resistance traits, and the diverse range of serotypes in the virulent DEC identified in this study confirm that differences exist between enteropathogens affecting children from interior towns of Uruguay and those circulating among children in Montevideo.

The Kinase Activity of Hematopoietic Progenitor Kinase 1 Is Essential for the Regulation of T Cell Function.

We examined hematopoietic protein kinase 1 (HPK1), whose reliance on scaffold versus kinase functions for negative immune cell regulation is poorly understood and critical to its assessment as a viable drug target. We identify kinase-dependent roles for HPK1 in CD8 T cells that restrict their anti-viral and anti-tumor responses by using HPK1 kinase-dead (HPK1.kd) knockin mice. Loss of HPK1 kinase function enhanced T cell receptor signaling and cytokine secretion in a T-cell-intrinsic manner. In response to chronic lymphocytic choriomeningitis virus (LCMV) infection or tumor challenge, viral clearance and tumor growth inhibition were enhanced in HPK1.kd mice, accompanied by an increase in effector CD8 T cell function. Co-blockade of PD-L1 further enhanced T effector cell function, resulting in superior anti-viral and anti-tumor immunity over single target blockade. These results identify the importance of HPK1 kinase activity in the negative regulation of CD8 effector functions, implicating its potential as a cancer immunotherapy target.

Tumour and host cell PD-L1 is required to mediate suppression of anti-tumour immunity in mice.

Expression of PD-L1, the ligand for T-cell inhibitory receptor PD-1, is one key immunosuppressive mechanism by which cancer avoids eradication by the immune system. Therapeutic use of blocking antibodies to PD-L1 or its receptor PD-1 has produced unparalleled, durable clinical responses, with highest likelihood of response seen in patients whose tumour or immune cells express PD-L1 before therapy. The significance of PD-L1 expression in each cell type has emerged as a central and controversial unknown in the clinical development of immunotherapeutics. Using genetic deletion in preclinical mouse models, here we show that PD-L1 from disparate cellular sources, including tumour cells, myeloid or other immune cells can similarly modulate the degree of cytotoxic T-cell function and activity in the tumour microenvironment. PD-L1 expression in both the host and tumour compartment contribute to immune suppression in a non-redundant fashion, suggesting that both sources could be predictive of sensitivity to therapeutic agents targeting the PD-L1/PD-1 axis.

Immunogenic peptide mimotopes from an epitope of Escherichia coli O157 LPS.

Escherichia coli O157:H7 is a subtype of Shiga toxin-producing E. coli that is associated with haemorrhagic colitis and haemolytic uremic syndrome (HUS). Studies of populations in endemic areas have reported that the presence of specific antibodies against the O157 lipopolysaccharide (LPS) is associated with a lower incidence of diarrhoea and HUS. Phage display and IgG anti-O157 LPS antibodies were used in the present study to select peptide mimotopes of O157 LPS expressed in protein III of the M13 phage. Synthetic peptides (SP) were designed using the derived amino acid sequences obtained from DNA nucleotides of 63 selected phagotopes. The LxP/YP/SxL motif was identified in five of the phagotope amino acid sequences. Antibody responses against the phagotopes and their corresponding SPs were evaluated. SP12, one of the designed SP, induced the production of antibodies against the homologous peptide (1:800) and O157 LPS (1:200). The specificity of anti-SP12 antiserum was confirmed by analyzing its response to SP3, an SP with a different amino acid sequence than that of SP12, as well as against an E. coli LPS different from O157. Competitive studies with SP12 and O157 LPS showed a significant decrease in anti-SP12 and anti-LPS O157 antiserum responses against SP12 and O157 LPS, respectively. Eighteen (82%) of the 22 human serum samples with positive reactivity against E coli O157 LPS reacted with SP12 SP (cut-off >0.4). These results support the idea that SP12 is an immunogenic mimotope of O157 LPS.

UPEC strain characterization isolated from Mexican patients with recurrent urinary infections.

INTRODUCTION: Any microorganism is capable of causing urinary tract infections (UTI). However, Uropathogenic Escherichia coli (UPEC) is responsible for the majority of UTI. A variety of virulence genes have been identified in UPEC. Diverse epidemiological studies support that specific subsets of genes are characteristic of each E. coli uropathogenic subtype involved in the development of cystitis, pyelonephritis, and urosepsis. METHODOLOGY: Twenty-three E. coli strains isolated from women with cystitis, 25 from men with prostatitis were characterized according to serotype, virulence genes, PFGE profile and susceptibility antimicrobials. RESULTS: E. coli O25:H4-ST131 was more frequently isolated from cystitis than prostatitis. The majority of cystitis strains presented almost all the virulence genes, contrary to 60% of prostatitis strains. The strains characterized by serology, PFGE and MLST and virulence profile showed that cystitis strains shared a lineage with their own genomic fingerprint, indicating that these strains derived from a common ancestor. The prostatitis strains showed serotypes diverse and much higher degree of genetic diversity, indicating that they are an unrelated group.  More than 50% of isolates were resistant to at least 7 of the antimicrobials tested. Antibiotic-tolerant cells were observed, presenting with more frequency in response chloramphenicol, tetracycline, trimethoprim-sulfometoxazole and dicloxicillin. CONCLUSIONS: Persister cells can be detected from the beginning of the infection. The importance of these persister cells that exhibit multidrug tolerance is that a single surviving bacterium can initiate again an infection making it clinical relevant in the chronic urinary tract infection.

Enteropathogens associated with acute diarrhea in children from households with high socioeconomic level in uruguay.

Infectious diarrhea, a common disease of children, deserves permanent monitoring in all social groups. To know the etiology and clinical manifestations of acute diarrhea in children up to 5 years of age from high socioeconomic level households, we conducted a descriptive, microbiological, and clinical study. Stools from 59 children with acute community-acquired diarrhea were examined, and their parents were interviewed concerning symptoms and signs. Rotavirus, adenovirus, and norovirus were detected by commercially available qualitative immunochromatographic lateral flow rapid tests. Salmonella, Campylobacter, Yersinia, and Shigella were investigated by standard bacteriological methods and diarrheagenic E. coli by PCR assays. We identified a potential enteric pathogen in 30 children. The most frequent causes of diarrhea were enteropathogenic E. coli (EPEC), viruses, Campylobacter, Salmonella, and Shiga-toxin-producing E. coli (STEC). Only 2 patients showed mixed infections. Our data suggest that children with viral or Campylobacter diarrhea were taken to the hospital earlier than those infected with EPEC. One child infected with STEC O26 developed "complete" HUS. The microbiological results highlight the importance of zoonotic bacteria such as atypical EPEC, Campylobacter, STEC, and Salmonella as pathogens associated with acute diarrhea in these children. The findings also reinforce our previous communications about the regional importance of non-O157 STEC strains in severe infant food-borne diseases.

Patrones de aceptación de terapia de hidratación oral a nivel domiciliario en una cohorte de madres rurales / Patterns of household acceptance of oral rehydration therapy in a cohort of rural mothers

Se estudió la aceptación de soluciones de hidratación oral a nivel domiciliario en cuadros subsecuentes de diarrea en una cohorte de 75 niños rurales seguidos longitudinalmente durante los primeros dos años de vida. El estudio se llevó a cabo entre agosto de 1985 y febrero de 1988 en un poblado rural del centro de México. Se detectaron 636 episodios de diarrea, con un promedio de cuatro episodios por niño por año. No hubo muertes por diarrea aguda y sus complicaciones y sólo el 2%de estos cuadros requirieron aplicación de soluciones parenterales. La proporción de madres que aceptaron administrar electrolitos orales en cuadros subsecuentes de diarrea fué constante alrededor del 50%. La aceptación de la terapia de hidratación oral se asoció con significancia estadística con un nivel socioeconómico medio-alto, la lectura de periódicos y revistas por la padre y la presencia de agua potable intradomiciliaria. Madres pertenecientes a familias clasificadas como tradicionales rechazaron con mayor frecuencia este tipo de tratamiento

Estudio comparativo entre el uso del método de Spicer-Edwards y la relación de coaglutinación para identificación de antígenos de Salmonella / Comparative study between Spicer-Edwards method and a coagglutination method to detect Salmonella antigen

Se realizó un estudio comparativo para la identificación de antígenos flagelares de Salmonella empleando el método de aglutinación en tubo con sueros de Spicer-Edwards y la reacción de coaglutinación utilizando proteína A de Staphylococcus aureus Cowan 1 (NCTC 8530). Se tipificaron por ambos métodos 39 serotipos de Salmonella pertenecientes a ocho serogrupos del esquema antigénico de Kauffman-White. Cada uno de los serogrupos incluía salmonelas de los diferentes serotipos monofásicos y bifásicos. El análisis estadístico de los resultados demostró que la reacción de coaglutinación tuvo mayor sensibilidad y especificidad para la detección de antígenos flagelares que la reacción clásica de aglutinación en tubo. El uso de menor cantidad de antisueros para detectar antígenos de Salmonella sin pérdida de sensibilidad y/o especificidad en la prueba de coaglutinación representa un ahorro importante para laboratorios clínios y de investigación epidemiológica que requieren de la identificación de estos microorganismos

Etiología de diarrea con sangre en niños de una comunidad rural / Etiology of bloody diarrhea in children of a rural village

Se investigó la etiología de diarrea con sangre en una cohorte de 75 niños rurales seguidos longitudinalmente durante los primeros dos años de vida. De un total de 636 episodios de diarrea, 71 (11%) mostraron la presencia de sangre. Se aisló un patógeno en 59 (83%) de los 71 casos; 35%mostraron la presencia de Escherichia coli enteroagregativa (EAEC), 11%de E. coli productora de toxinas tipo Shiga (SLT) I y/o II; 13%de Shigella, 7%de Campylobacter jejuni, o de E. coli enterotoxigénica (ETEC); y en 4%de E. coli enteropatógena (EPEC), E. coli enteroinvasora, Salmonella, Giardia lamblia, o Hymenolepis nana. Se encontraron cultivos mixtos en 16%de casos durante el primer año y en 18%durante el segundo. Estas mezclas fueron principalmente combinaciones de C. jejuni o ETEC con EAEC. el 78%de los niños con diarrea con sangre cursaron con un solo evento diarreico durante los primeros dos años de vida con duración promedio de siete días. Epidemiológicamente, la incidencia de diarrea con sangre se incrementó progresivamente durante los primeros dos años de vida, con un descenso significativo después de los diez meses de edad. La prevalenia de patógenos estuvo afectada por la estacionalidad, excepto para EAEC cuya prevalencia fué endémica. En el caso de Shigella y E. coli productora de SLT, la agrupación de casos en tiempo y el aislamiento de especies serológicamente idénticas indicaron que las infecciones fueron resultado de brotes de una fuente común

Producción de citotoxina tipo shiga (SLT) 1 en cepas de Escherichia coli aisladas de niños con diarrea en una comunidad rural / Production of shiga like toxin (SLT) 1 in strains of Escherichia coli isolated from infants with diarrhoea in arural community

Se estudió la producción de citotoxina tipo shiga-1 (SLT) por la prueba de neutralización descrita por O'Brien y col. en 1,006 cepas de Escherichia coli aisladas de 52 niños con diarrea y de 52 controles apareados por edad, sexo y vecindad, pertenecientes todos a una cohorte de niños rurales, la cual ha sido seguida longitudinalmente durante los primeros dos años de vida a partir del nacimiento. Se buscó la producción de enterotoxinas estable y lábil al calentamiento y la presencia de factores de colonización I, II y E8775 en las mismas bacterias, así como la habilidad para adherirse a células HEp-2 y de aglutinar contra antisueros pertenecientes a los serogrupos enteropatógenos de Escherichia coli, incluyendo 0142 y 0157. Los niños con diarrea presentaron aislamiento significativamente mayor de cepas productoras de citotoxina en cantidades moderadas y altas (p < 0.01). No se encontró relación entre producción de citotoxina y otras características de patogenicidad en cepas aisladas en ambos grupos. El 30% de las bacterias productoras de cantidades moderadas de citotoxina y el 80% de las productoras de cantidades moderadas elevadas pertenecieron al serotipo 0157:H7. En tres niños con diarrea y dos sin diarrea se aislaron cepas productoras de otra citotoxina cuyo efecto no fue neutralizable con un antisuero específico contra SLT-1. Estas cepas han sido confirmadas como productoras de SLT-2. Escherichia coli enterohemorrágica, citotoxina tipo Shiga; diarrea; colitis hemorrágica afebril