Resident training in interventional pathology: Ultrasound-guided fine-needle aspiration and rapid on site evaluation-5 years of teaching experience in a single university hospital.

INTRODUCTION: Despite the established role of the interventional pathologist, their diagnostic performance is difficult to establish. At least in Spain training of pathology residents in ultrasound-guided interventional procedures for specimen collection is limited or absent in most institutions. We present our teaching experience in the instruction of ultrasound-guided fine-needle aspiration (FNA) to pathology residents in a tertiary-level hospital. MATERIALS AND METHODS: The training of pathology residents who rotated through the interventional unit of the pathology department and the application of ultrasound-guided FNA and rapid on-site evaluation (U-ROSE) was documented over 5 years. The training period was broken down into learning phases and included the number of ultrasound-guided FNA performed, anatomical location, and their diagnostic performance, among other aspects. RESULTS: Nineteen (19) pathology residents were trained in U-ROSE, and performed a total of 4003 procedures, with a mean of 211 per resident. In 53% of cases only one pass was required for an adequated sample. The specimen was diagnostic in more than 97% of cases. The most frequently sampled anatomical sites were the thyroid gland (n = 2347), followed by lymph node (n = 667), soft tissues (n = 663) and salivary glands (n = 322). CONCLUSION: The results support the training programme followed by pathology residents in learning U-ROSE, which is essential to lay the foundations for the future interventional pathologist.

Intranasal challenge with B. pertussis leads to more severe disease manifestations in mice than aerosol challenge.

The murine Bordetella pertussis challenge model has been utilized in preclinical research for decades. Currently, inconsistent methodologies are employed by researchers across the globe, making it difficult to compare findings. The objective of this work was to utilize the CD-1 mouse model with two routes of challenge, intranasal and aerosol administration of B. pertussis, to understand the differences in disease manifestation elicited via each route. We observed that both routes of B. pertussis challenge result in dose-dependent colonization of the respiratory tract, but overall, intranasal challenge led to higher bacterial burden in the nasal lavage, trachea, and lung. Furthermore, high dose intranasal challenge results in induction of leukocytosis and pro-inflammatory cytokine responses compared to aerosol challenge. These data highlight crucial differences in B. pertussis challenge routes that should be considered during experimental design.

Pleural metastasis from pulmonary primary signet-ring squamous cell carcinoma. Case report and literature review.

Signet-ring cells are morphologically defined by the presence of a large intracytoplasmic vacuole that compresses and displaces the nucleus to the periphery. In most cases, these cells are associated with adenocarcinomas of various locations, and with non-epithelial neoplasms. To date, less than 20 cases of squamous cell carcinoma with signet-ring morphology have been described, mainly located on the skin. We present the case of a 73-year-old male with pleural effusion and a left lower lobe mass. The cytological study of the pleural effusion allowed the diagnosis of metastasis of squamous cell carcinoma, signet-ring cell variant. The treatment of lung cancer in advanced stages requires a precise diagnosis that allows the best therapy to be offered to the patient, depending on the clinical stage and the positivity of the biomarkers, among others. Our patient died 18 months after the initial diagnosis.

Cyclic di-GMP Regulates the Type III Secretion System and Virulence in Bordetella bronchiseptica.

The second messenger cyclic di-GMP (c-di-GMP) is a ubiquitous molecule in bacteria that regulates diverse phenotypes. Among them, motility and biofilm formation are the most studied. Furthermore, c-di-GMP has been suggested to regulate virulence factors, making it important for pathogenesis. Previously, we reported that c-di-GMP regulates biofilm formation and swimming motility in Bordetella bronchiseptica. Here, we present a multi-omics approach for the study of B. bronchiseptica strains expressing different cytoplasmic c-di-GMP levels, including transcriptome sequencing (RNA-seq) and shotgun proteomics with label-free quantification. We detected 64 proteins significantly up- or downregulated in either low or high c-di-GMP levels and 358 genes differentially expressed between strains with high c-di-GMP levels and the wild-type strain. Among them, we found genes for stress-related proteins, genes for nitrogen metabolism enzymes, phage-related genes, and virulence factor genes. Interestingly, we observed that a virulence factor like the type III secretion system (TTSS) was regulated by c-di-GMP. B. bronchiseptica with high c-di-GMP levels showed significantly lower levels of TTSS components like Bsp22, BopN, and Bcr4. These findings were confirmed by independent methods, such as quantitative reverse transcription-PCR (q-RT-PCR) and Western blotting. Higher intracellular levels of c-di-GMP correlated with an impaired capacity to induce cytotoxicity in a eukaryotic cell in vitro and with attenuated virulence in a murine model. This work presents data that support the role that the second messenger c-di-GMP plays in the pathogenesis of Bordetella.

Ultrasound-Guided Fine-Needle Aspiration of Superficial Lymphadenopathy Performed by Interventional Pathologists: The Applicability of the Sydney System from 2 Years of Experience and 363 Cases.

INTRODUCTION: The Sydney system proposal for the study and reporting of lymphadenopathy by fine-needle aspiration (FNA) constitutes one of the first attempts to standardize this procedure. Here, we review its applicability. MATERIALS AND METHODS: A retrospective study in which all ultrasound-guided FNAs (USFNAs) of superficial lymphadenopathy (palpable or not) performed by interventional pathologists in 2 specialized hospital centers were quantified over 2 years. The procedure was systematized, and the diagnoses were reclassified according to the Sydney system categories. RESULTS: We analyzed 363 USFNAs of lymphadenopathies. The distribution of cases by categories was as follows: insufficient (n = 13; 3.58%), benign (n = 208; 57.30%), atypia of uncertain significance (n = 7; 1.93%), suspicious (n = 21; 5.79), and malignant (n = 114; 31.40%). The risks of malignancy calculated for categories I, II, III, IV, and V were 27%, 3%, 50%, 100%, and 100%, respectively. CONCLUSION: The implementation of the Sydney system allows the systematization and standardization of the lymph node FNA methodology, with increased efficacy and efficiency. Assimilating the recommendations enables the qualification of the diagnostic procedure.

[Minimally invasive percutaneous musculoskeletal biopsy performed by Interventional Pathologist for myopathy study. Twenty years of experience]. / Biopsia percutánea muscular mínimamente invasiva realizada por patólogo intervencionista en el estudio de miopatías. Veinte años de experiencia.

INTRODUCTION: Muscle biopsy plays a major role in the final diagnosis of myopathies. Open muscle biopsy is the benchmark procedure, although minimally invasive percutaneous muscle biopsy (MIPMB) has demonstrated comparable diagnostic performance at a lower cost and can be carried out by interventional pathologists. MATERIALS AND METHODS: Muscle biopsies performed from 1997 to 2017 were reviewed and classified according to the type of procedure, whether carried out by an interventional pathologist or another specialist, the diagnosis and the effectiveness of the procedure. RESULTS: 738 muscle biopsies were performed; 32% were open biopsies and 68% MIPMB carried out by pathologist. The muscle most often biopsied was the femoral quadriceps and the most frequent diagnosis was inflammatory myopathies. In only 39 cases (20 open biopsies and 19 MIPMB) was there insufficient tissue for diagnosis. CONCLUSIONS: Muscle biopsy proved highly effective as a diagnostic tool as 90% yielded adequate tissue samples. The results obtained with MIPMB performed by interventionist pathologists were comparable to those of open muscle biopsy.

Intranasal acellular pertussis vaccine provides mucosal immunity and protects mice from Bordetella pertussis.

Current acellular pertussis vaccines fall short of optimal protection against the human respiratory pathogen Bordetella pertussis resulting in increased incidence of a previously controlled vaccine- preventable disease. Natural infection is known to induce a protective mucosal immunity. Therefore, in this study, we aimed to use acellular pertussis vaccines to recapitulate these mucosal immune responses. We utilized a murine immunization and challenge model to characterize the efficacy of intranasal immunization (IN) with DTaP vaccine or DTaP vaccine supplemented with curdlan, a known Th1/Th17 promoting adjuvant. Protection from IN delivered DTaP was compared to protection mediated by intraperitoneal injection of DTaP and whole-cell pertussis vaccines. We tracked fluorescently labeled DTaP after immunization and detected that DTaP localized preferentially in the lungs while DTaP with curdlan was predominantly in the nasal turbinates. IN immunization with DTaP, with or without curdlan adjuvant, resulted in anti-B. pertussis and anti-pertussis toxin IgG titers at the same level as intraperitoneally administered DTaP. IN immunization was able to protect against B. pertussis challenge and we observed decreased pulmonary pro-inflammatory cytokines, neutrophil infiltrates in the lung, and bacterial burden in the upper and lower respiratory tract at day 3 post challenge. Furthermore, IN immunization with DTaP triggered mucosal immune responses such as production of B. pertussis-specific IgA, and increased IL-17A. Together, the induction of a mucosal immune response and humoral antibody-mediated protection associated with an IN administered DTaP and curdlan adjuvant warrant further exploration as a pertussis vaccine candidate formulation.

Bordetella bronchiseptica Diguanylate Cyclase BdcA Regulates Motility and Is Important for the Establishment of Respiratory Infection in Mice.

Bacteria can be motile and planktonic or, alternatively, sessile and participating in the biofilm mode of growth. The transition between these lifestyles can be regulated by a second messenger, cyclic dimeric GMP (c-di-GMP). High intracellular c-di-GMP concentration correlates with biofilm formation and motility inhibition in most bacteria, including Bordetella bronchiseptica, which causes respiratory tract infections in mammals and forms biofilms in infected mice. We previously described the diguanylate cyclase BdcA as involved in c-di-GMP synthesis and motility regulation in B. bronchiseptica; here, we further describe the mechanism whereby BdcA is able to regulate motility and biofilm formation. Amino acid replacement of GGDEF with GGAAF in BdcA is consistent with the conclusion that diguanylate cyclase activity is necessary for biofilm formation and motility regulation, although we were unable to confirm the stability of the mutant protein. In the absence of the bdcA gene, B. bronchiseptica showed enhanced motility, strengthening the hypothesis that BdcA regulates motility in B. bronchiseptica We showed that c-di-GMP-mediated motility inhibition involved regulation of flagellin expression, as high c-di-GMP levels achieved by expressing BdcA significantly reduced the level of flagellin protein. We also demonstrated that protein BB2109 is necessary for BdcA activity, motility inhibition, and biofilm formation. Finally, absence of the bdcA gene affected bacterial infection, implicating BdcA-regulated functions as important for bacterium-host interactions. This work supports the role of c-di-GMP in biofilm formation and motility regulation in B. bronchiseptica, as well as its impact on pathogenesis.IMPORTANCE Pathogenesis of Bordetella spp., like that of a number of other pathogens, involves biofilm formation. Biofilms increase tolerance to biotic and abiotic factors and are proposed as reservoirs of microbes for transmission to other organs (trachea, lungs) or other hosts. Bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) is a second messenger that regulates transition between biofilm and planktonic lifestyles. In Bordetella bronchiseptica, high c-di-GMP levels inhibit motility and favor biofilm formation. In the present work, we characterized a B. bronchiseptica diguanylate cyclase, BdcA, which regulates motility and biofilm formation and affects the ability of B. bronchiseptica to colonize the murine respiratory tract. These results provide us with a better understanding of how B. bronchiseptica can infect a host.