Phenotypic and genotypic characterization of enteroaggregative Escherichia coli isolates from pediatric population in Pakistan

Enteroaggregative Escherichia coli (EAEC) are a leading cause of diarrhea among children. The objective of this study was to define the frequency of EAEC among diarrheal children from flood‐affected areas as well as sporadic cases, determine multidrug resistance, and evaluation of virulence using an in vivo model of pathogenesis. Stool samples were collected from 225 diarrheal children from 2010 to 2011 from flood‐affected areas as well as from sporadic cases in Pakistan. Identified EAEC isolates were characterized by phylogrouping, antibiotic resistance patterns including the extended‐spectrum beta lactamase spectrum, single nucleotide polymorphism detection in gyrA and parC, and virulence potential using wax worm, G. mellonella. A total of 35 (12.5%) confirmed EAEC isolates were identified among 225 E. coli isolates. EAEC isolates displayed high resistance to tetracycline, ampicillin, and cefaclor. A total of 34.28% were ESBL positive. Single nucleotide polymorphism detection revealed 37.14% and 68.57% isolates were positive for SNPs in gyrA (A₆₆₀‐T₆₆₀) and parC (C₃₃₀‐T₃₃₀), respectively. Phylogrouping revealed that B2 phylogroup was more prevalent among all EAEC isolates tested followed by D, A, B1, and non‐typeable (NT). Infection of G. mellonella with EAEC showed that killing infective dose was 100% higher than E. coli DH5 alpha control. EAEC are prevalent among Pakistani children with diarrhea, they are highly resistant to antibiotics, and predominantly fall into B2 phylogroup. Epidemiologic surveillance of EAEC and other E. coli pathotypes is critical to assess not only the role of these pathogens in diarrheal disease but also to determine the extent of multidrug resistance among the population.     

Fine-needle aspiration biopsy of cystic pleomorphic adenoma with adnexa-like differentiation mimicking mucoepidermoid carcinoma: a case report

Although pleomorphic adenomas (PAs) usually can be diagnosed very accurately with fine-needle aspiration biopsy (FNAB), even this common salivary gland neoplasm can be diagnostically challenging and cause pitfalls in cytodiagnosis. In particular, the presence of either cystic changes or squamous, mucinous, or sebaceous metaplasia can lead to a false positive diagnosis of mucoepidermoid carcinoma (MEC). Here, we present a case of a 70-yr-old man with an asymptomatic left deep lobe parotid mass for which CT-guided FNAB was performed. The FNAB cytology revealed cohesive clusters of squamous epithelial cells, sebaceous cells, oncocytes, macrophages, and rare myoepithelial cells. Characteristic metachromatic fibrillar chondromyxoid stroma, which usually is seen in PAs, was not seen in the aspirate. Although cytodiagnosis of PAs was suggested based on the presence of other cellular components, resection was recommended. The subsequent parotidectomy specimen revealed an encapsulated cystic PA with mixed appendageal differentiation including areas of squamous, mucinous, sebaceous, and oncocytic metaplasia. Chondromyxoid stroma was only focally present. Presence of squamous, mucinous, and/or sebaceous metaplasia, especially in the absence of chondromyxoid stroma, presents the potential for misinterpretation of the FNAB as indicative of malignancy in general and MEC in particular.     

Role of La autoantigen and polypyrimidine tract-binding protein in HCV replication

To determine if the cellular factors La autoantigen (La) and polypyrimidine tract-binding protein (PTB) are required for hepatitis C virus (HCV) replication, we used siRNAs to silence these factors and then monitored their effect on HCV replication using quantitative RT-PCR. In addition, we determined the influence of PTB on the activity of the 3' noncoding region (NCR) of HCV and investigated its interaction with the components of the HCV replicase complex. We found that La is essential for efficient HCV replication while PTB appears to partially repress replication. PTB does, however, block the binding of HCV RNA-dependent RNA polymerase (RdRp, NS5B) to the 3'NCR. Indirect immunofluorescence microscopy showed co-localization of cytoplasmic PTB with the HCV RdRp in hepatoma cells (Huh-7) expressing HCV proteins, while in vitro translation of viral proteins from the HCV replicon revealed the interaction of PTB isoforms with NS5B polymerase and NS3.     

Estrogen receptor (ER), progesterone receptor (PR), and HER2 expression pre- and post- neoadjuvant chemotherapy in primary breast carcinoma: a single institutional experience

Background

The estrogen receptor (ER), progesterone receptor (PR), and HER2 profile of a primary breast carcinoma plays a significant role in patient management and treatment. Because of the increasing utilization of neoadjuvant chemotherapy or hormone therapy, surgically-resected carcinomas often show marked treatment effect. The aim of this study was to compare immunohistochemical (IHC) profiles (ER, PR, HER2, HER2 FISH) of primary breast carcinomas before and after neoadjuvant chemotherapy to assess the subsequent effects on hormone receptor status.

Design

Primary breast carcinomas from 38 female patients treated with neoadjuvant therapy after needle core biopsy or fine needle aspiration diagnosis were included. Histologic data was collected for each case, including site, type, grade, tumor size (cm), pre- and post- neoadjuvant treatment IHC panel (ER, PR, HER2), and fluorescence in-situ hybridization (FISH) for HER2.

Results

Of the 38 carcinomas studied, 45 % were positive for ER by IHC both pre- and post- neoadjuvant treatment (P=1.00). IHC studies for PR in these 38 patients showed 37% positivity for PR pre-neoadjuvant therapy and 21% positivity post-treatment (p=0.03). For 37 patients with HER2 IHC, 32% were positive pre-treatment, and 22% were positive post-treatment (P = 0.20). For 7 patients, HER2 FISH was positive in 71% pre-therapy and in 57% post-treatment (P=0.32).

Conclusions

Profiles for ER, HER2 IHC, and HER2 FISH were not significantly different in primary breast carcinomas before and after neoadjuvant chemotherapy. Further investigation is warranted to assess reproducibility of technique and investigate clinical implications of significant loss of PR status in treated patients.     

Dose-dependent effect of histamine on liver function markers in immunized rabbits

ObjectiveThe present study was designed to delineate the hepatotoxicological roles of histamine dose-dependently in immunized rabbits.MethodsThe cohort comprised of three groups (II, III and IV), containing 18 rabbits each, and received subcutaneous histamine 50 μg/kg, 100 μg/kg and 200 μg/kg, respectively for 10 days (b.i.d., starting from 3 days prior to immunization until 7 days after immunization). Group I (control, n = 18) received subcutaneous sterile distilled water for 10 days. They were subsequently immunized at day 3 with intravenous injection of SRBC (1 × 10⁹ cells/ml). Blood samples were collected on pre-immunization (pre-I) day 0, as well as on days 7-, 14-, 21-, 28- and 58-post-immunization (post-I). Biochemical parameters aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP) and bilirubin [total bilirubin (TB), direct bilirubin (DB) and indirect bilirubin (IB)] were determined.ResultsGroups II and IV revealed a significant decrease (on day 0-pre-I) and a significant increase (on days 7-, 14-, 21-, 28- and 58-post-I) in ALT and AST levels, when compared with the corresponding values of groups I and III while group II showed a significant increase in ALT and AST levels as compared to group IV. ALP levels in groups II, III and IV showed a significant enhancement when compared with group I. Moreover, results of TB, DB and IB demonstrated increased levels in group III when compared with groups I, II and IV. The results were found statistically significant (p < 0.05).ConclusionShort-term treatment of histamine produces dose-dependent differential patterns of hepatic dysfunctions suggestive mild liver degeneration warranting further long-term studies.     

Fascin stain as a potential marker of invasiveness in carcinomas of the urinary bladder: a retrospective study with biopsy and cytology correlation

The evaluation of invasion in urothelial carcinomas of the urinary bladder cannot be determined on cytology and can be particularly challenging in biopsy cases with limited sampling. Recent studies of bladder resection specimens suggest that fascin overexpression may be a marker of aggressive urothelial carcinomas and can help facilitate the assessment of invasion. In this study, we evaluated urine cytology and corresponding biopsy specimens with proven invasive urothelial carcinoma for fascin expression by immunohistochemistry. Thirty‐five patients diagnosed with positive urine cytology and biopsy‐proven invasive urothelial carcinoma between January 2003 and February 2009 were identified. We found increased fascin expression in 100% (35/35) of SurePathTM&!trade; urine cytology preparations as well as 100% (35/35) of corresponding biopsy cases with invasive urothelial carcinoma. On urine cytology, cytoplasmic fascin staining was moderate to intense in malignant tumor cell clusters and single cells and not observed in benign urothelial cells. Staining in biopsy cases was generally intense and cytoplasmic and present in both the invasive (100%) and noninvasive (31%) components of the lesion. These findings uphold the association of increased fascin expression in invasive urothelial carcinomas of the urinary bladder. We furthermore demonstrate that fascin staining can be performed successfully on SurePathTM&!trade; urine cytology preparations in which increased fascin expression correlates with invasion on biopsy. While not a definitive marker of invasion, as it is observed in in situ carcinoma, we conclude that the utilization of fascin immunohistochemistry on urine cytology might serve as a useful adjunct in predicting invasiveness in subsequent biopsies. Diagn. Cytopathol. 2010. © 2010 Wiley‐Liss, Inc.     

TLR engagement prior to virus infection influences MHC-I antigen presentation in an epitope-dependent manner as a result of nitric oxide release

Microorganisms contain PAMPs that can interact with different TLR-Ls. Cooperative signals from these receptors may modify innate and adaptive immune responses to invading pathogens. Therefore, a better understanding of the role TLRs play in initiating host defense during infections requires assessing the influence of multiple TLR engagement on pAPC activation and antigen presentation. In this study, we investigated the effects of combined TLR2, TLR3, or TLR4 engagement on DC activation and the presentation of LCMV antigens focusing on the major epitopes derived from NP and GP proteins encoded by the virus. Our results demonstrate that combined TLR ligation affected antigen presentation of NP(205-212), GP(33-41), and GP(276-286), but not NP(396-404). The altered antigen presentation was associated with changes in proteasomal activities and NO production as a result of TLR engagement. Taken together, the data demonstrate that combined TLR ligation could result in changes of innate effectors that may directly influence the adaptive immune response.     

Fine‐needle aspiration cytology diagnosis of non‐Hodgkins lymphoma in a resource‐challenged environment

To establish the role of fine‐needle aspiration cytology (FNAC) as a diagnostic tool for non‐Hodgkins lymphoma in a resource challenged environment. This study was conducted on patients with lymphadenopathy, attending various clinics over a period of 18 months. FNAC of the enlarged lymph nodes was performed and biopsy, special stains and immunohistochemical staining was done in selected cases. Out of the total 275 cases, 42 cases (16%) were primary lymphoproliferative disorders. Non‐Hodgkin lymphoma comprised of 32 cases (76.2% of all lymphomas), Hodgkin lymphoma‐10 cases and the rest were metastatic carcinoma. The diagnostic accuracy for non‐Hodgkin Lymphoma was 93.3%, sensitivity 95.4%, and specificity 87.5%. FNAC is a rapid, safe, easy, and nonexpensive diagnostic technique which can be used for early diagnosis of non‐Hodgkins lymphoma. Diagn. Cytopathol. 2011. © 2010 Wiley‐Liss, Inc.