Diagnosis of Hirschsprung disease by analyzing acetylcholinesterase staining using artificial intelligence.

OBJECTIVES: Classical Hirschsprung disease (HD) is defined by the absence of ganglion cells in the rectosigmoid colon. The diagnosis is made from rectal biopsy, which reveals the aganglionosis and the presence of cholinergic hyperinnervation. However, depending on the method of rectal biopsy, the quality of the specimens and the related diagnostic accuracy varies substantially. To facilitate and objectify the diagnosis of HD, we investigated whether software-based identification of cholinergic hyperinnervation in digitalized histopathology slides is suitable for distinguishing healthy individuals from affected individuals. METHODS: N = 190 samples of 112 patients who underwent open surgical rectal biopsy at our pediatric surgery center between 2009 and 2019 were included in this study. Acetylcholinesterase (AChE) stained slides of these samples were collected and digitalized via slide scanning and analyzed using two digital imaging software programs (HALO, QuPath). The AChE-positive staining area in the mucosal layers of the intestinal wall was determined. In the next step machine learning was employed to identify patterns of cholinergic hyperinnervation. RESULTS: The area of AChE-positive staining was greater in HD patients compared to healthy individuals (p < 0.0001). Artificial intelligence-based assessment of parasympathetic hyperinnervation identified HD with a high precision (area under the curve [AUC] 0.96). The accuracy of the prediction model increased when nonrectal samples were excluded (AUC 0.993). CONCLUSIONS: Software-assisted machine-learning analysis of AChE staining is suitable to improve the diagnostic accuracy of HD.

Clinical relevance of inflammation on rectal biopsy for Hirschsprung disease: An outcomes analysis.

OBJECTIVES: Inflammation on diagnostic rectal biopsy for children with suspected Hirschsprung disease (HSCR) is reported on pathology, and its significance is unknown. We describe the management and outcomes of a cohort with inflammation on rectal biopsy compared to those without. Specifically, to address the hypothesis that inflammation on diagnostic biopsy is associated with increased complication rates irrespective of intervention type and timing. METHODS: A single institution retrospective review of children with HSCR who underwent biopsy and endorectal pull-through (ERPT) from 2010 to 2020 was performed. The primary outcome was overall complications at 30-days following ERPT. Secondary outcomes included timing and type of operative intervention as well as postoperative enterocolitis diagnosed within 6-months of ERPT. RESULTS: Forty-nine children were identified; inflammation was present on diagnostic biopsy for 17 children. Those with inflammation were more likely to have clinical evidence of enterocolitis at the time of biopsy (p = 0.001) and were more likely to undergo leveling colostomy before ERPT (p = 0.01). Children with inflammation had a higher anastomotic leak rate (p = 0.04). Subgroup analysis of patients with inflammation undergoing primary ERPT versus leveling colostomy demonstrated no significant difference in outcomes following definitive ERPT. CONCLUSIONS: Our study suggests inflammation on diagnostic rectal biopsy for HSCR is associated with increased anastomotic leak rates. While additional prospective studies are indicated, attention to methods of mitigating inflammation and confirming its resolution before definitive pull-through may be of benefit for improving clinical outcomes in patients found with inflammation on diagnostic rectal biopsy.

Evaluation of diagnostic factors used to refer children with constipation for rectal biopsies.

PURPOSE: Children with constipation and suspected Hirschsprung's disease are referred for rectal biopsy. Since this is an invasive procedure, appropriate indications should be applied to minimize the number of "unnecessary" biopsies. METHODS: We reviewed all constipated children who underwent a rectal biopsy to diagnose a possible Hirschsprung's disease at a tertiary referral hospital over a 6-year period (2013-2018). We registered clinical and demographic factors in these children and conducted correlation and multivariate regression analysis to evaluate the relation between these factors and a diagnosis of Hirschsprung's disease. RESULTS: We identified 225 children, aged 0-17 years. In total, Hirschsprung's disease was diagnosed in only 49/225 (22%). Among the 49 children with Hirschsprung's disease, 29 (59%) were diagnosed in the neonatal period. Among girls, HD was confirmed in only 10/101 (10%) children, and only 1 of these 10 girls was older than 6 months at the time of the biopsy. The following factors correlated significantly with Hirschsprung's disease diagnosis in children older than 1 month: "male sex", "failure to thrive", "gross abdominal distention plus vomiting" and "fulfils the Rome 4 criteria for functional constipation". CONCLUSION: In children referred for rectal biopsy, the factors most indicative of Hirschsprung's disease were "male sex", "failure to thrive", "gross abdominal distention plus vomiting" and "fulfils the Rome 4 criteria for functional constipation". Notably, the prevalence of Hirschsprung's disease decreased with the increasing age of the children. Girls referred for a biopsy rarely had Hirschsprung's disease, especially those older than 1 month.

Calretinin Staining in Anorectal Line Biopsies Accurately Distinguished Hirschsprung Disease in a Retrospective Study.

INTRODUCTION: The absence of submucosal ganglion cells does not reliably distinguish Hirschsprung disease from non Hirschsprung disease in anorectal line biopsies. Calretinin staining might be helpful in these biopsies. To determine its value, we analyzed calretinin positive mucosal neurites in anorectal line biopsies. METHODS: Two pediatric pathologists, without access to patient data, evaluated calretinin positive mucosal neurites in anorectal line junctional mucosa in archival rectal biopsies contributed by 17 institutions. A separate investigator compiled patient information and sent data for statistical analysis. RESULTS: Biopsies with anorectal junctional mucosa from 115 patients were evaluated for calretinin positive mucosal neurites. 20/20 Hirschsprung disease biopsies were negative. 87/88 non Hirschsprung disease biopsies and 7/7 post pullthrough Hirschsprung disease neorectal biopsies were positive. Statistical analysis of the 108 non pullthrough biopsies yielded an accuracy of 99.1% (sensitivity 100%, specificity 98.9%). Age range was preterm to 16 years. Biopsy size was less than 1 mm to over 1 cm. CONCLUSIONS: Absence of calretinin positive mucosal neurites at the anorectal line was highly accurate in distinguishing Hirschsprung disease from non Hirschsprung disease cases in this blinded retrospective study. Calretinin staining is useful for interpreting biopsies from the physiologic hypoganglionic zone up to the anorectal line.

Rectal suction biopsy versus incisional rectal biopsy in the diagnosis of Hirschsprung disease.

BACKGROUND: Hirschsprung disease is one of the most common congenital anomalies that affect colorectal function. Rectal biopsy demonstrating the absence of ganglion cells in the affected bowel is the gold standard for diagnosis. Suction and incisional rectal biopsies are appropriate methods for obtaining diagnostic tissue. The goal of this study is to determine if any differences in adequacy exist between suction and incisional rectal biopsies at our institution. METHODS: We conducted a retrospective review of suction and incisional rectal biopsies for inadequacy per procedure at a tertiary pediatric hospital. Each procedure for rectal biopsy was also evaluated by a number of biopsies per procedure. We used a two-sample test of proportions to compare the inadequacy of suction vs. incisional biopsies. RESULTS: 133 rectal suction biopsy procedures (227 biopsies) and 125 incisional biopsy procedures (140 biopsies) were analyzed. In patients 6 months of age and older, the percentage of inadequate procedures was substantially higher in the suction biopsy group (24.1% vs 0.9%, p < 0.01). CONCLUSIONS: A substantially higher proportion of inadequacy was found in the suction rectal biopsy group compared to the incisional cohort among the older patient cohort, suggesting incisional biopsies should be strongly considered as the primary rectal biopsy method in patients older than 6 months.

Diagnostic accuracy of haematoxylin-eosin staining in comparison to calretinin and S100 for the assessment of ganglion cells in rectal biopsy.

OBJECTIVE: To assess the diagnostic accuracy of haematoxylin-eosin staining in clinically suspected Hirschsprung disease, and to compare the findings with calretinin and S100 immunohistochemistry. METHODS: The retrospective study was conducted at the AL-Khansaa Teaching Hospital, Nineveh, Iraq, and comprised data from January 2017 to October 2020 of rectal suction biopsies of patients with clinically and radiologically suspected Hirschsprung disease. Histopathology and immunohistochemistry were performed. Data was analysed using SPSS 16. RESULTS: Of the 114 patients, 74(64.9%) were males and 40(35.1%) were females. Based on histology, 28(24.6%) cases were negative for ganglion cells, and, of them 25(89.2%) revealed nerve bundle hypertrophy. The diagnostic accuracy for the detection of ganglion cell and nerve hypertrophy using haematoxylin-eosin stain was 99.1% and 94.4%, respectively. Correlation of haematoxylin-eosin staining with calretinin and S100 was statistically near perfection (κ= 0.976 and κ = 0.923), respectively. CONCLUSIONS: The mainstay to confirm or exclude Hirschsprung disease remains an accurate histopathological evaluation of the haematoxylin-eosin-stained sections of an adequate colorectal biopsy.

Gag p24 Is a Marker of Human Immunodeficiency Virus Expression in Tissues and Correlates With Immune Response.

We demonstrate that human immunodeficiency virus (HIV) gag p24 protein is more readily detected in gut and lymph node tissues than in blood CD4+ T cells and correlates better with CD4 count during antiretroviral therapy (ART). Gut p24 levels also measurably decline with ART in natural controllers. During ART, gut p24 expression is more strongly associated both with HIV-specific CD8+ T-cell frequency and plasma soluble CD14 levels than gut HIV RNA expression. This study supports using gag p24 as a marker of HIV expression in HIV+ tissues to study effects of viral persistence and to monitor efficacy of treatment in HIV-based clearance studies.

Autopsy Study of Calretinin Immunohistochemistry in the Anorectal Canal in Young Infants and Potential Implications for Rectal Biopsy Approach in the Neonatal Period.

BACKGROUND: Absent submucosal ganglion cells in biopsies 1-3 cm above the pectinate line establishes the pathologic diagnosis of Hirschsprung Disease (HD). Calretinin stains both ganglion cells and their mucosal neurites and has gained importance in HD diagnosis. Absent calretinin positive mucosal neurites in biopsies at the appropriate level above the pectinate line is highly specific for HD. Whether this applies to lower biopsies is uncertain. To address this, we studied anorectal canal autopsy specimens from infants. METHODS: We performed an autopsy study of infant anorectal canal specimens to describe calretinin staining in this region. Calretinin staining was correlated with histologic and gross landmarks. RESULTS: In all 15 non-HD specimens, calretinin positive mucosal neurites were present in glandular mucosa up to the anorectal line where neurites rapidly diminished. Age range was preterm 26 weeks to 3 months. CONCLUSIONS: Calretinin positive mucosal neurites are present in glandular mucosa up to the anorectal line in young infants. This is potentially important regarding neonatal HD biopsy level and diagnosis. Positive calretinin staining at the anorectal line favors normal innervation making HD unlikely. Absent calretinin positive neurites in glandular mucosa is worrisome for HD in young infants, regardless of location.

Acetylcholinesterase staining for the pathological diagnosis of Hirschsprung's disease.

Hirschsprung's disease (HD) is a congenital disease manifesting various degrees of functional bowel obstruction caused by the absence of enteric ganglion cells, which are usually absent in the colonic segment of the HD patient. Because the aganglionic segment of HD always includes the rectum, pathological diagnosis can be made using a rectal sample. HD should be diagnosed as early as possible because serious complications, such as acute enterocolitis or toxic megacolon, can develop without a definitive diagnosis and appropriate treatment. In the mid-1900s, HD was diagnosed by HE staining of specimens obtained by full-thickness biopsy. Since then, the combination of rectal mucosal biopsy and rubeanic acid-amplificated AChE staining has been brought about by the following milestones: the discovery that the submucosal plexus and the intermuscular plexus had the same level of nerve migration; the findings of research on acetylcholine (ACh) and acetylcholinesterase (AChE) in the intestinal tract; and the establishment of a rubeanic acid amplification method. Consequently, the diagnostic rate of HD improved dramatically in the 1980s. This review outlines the history of diagnostic methods for HD, the roles of ACh and AChE in the intestine, and the method of AChE staining.

Current practice of rectal biopsies for the diagnosis of Hirschsprung's disease in Latin America: an international online survey.

PURPOSE: The gold standard for the diagnosis of Hirschsprung's disease (HD) is a rectal biopsy. The sample may be obtained using a transanal excisional biopsy (TEB) or suction technique. Rectal suction biopsy (RSB) is not a standard procedure in Latin-America. AIM: To evaluate the current practice in rectal biopsy for HD among pediatric surgeons. METHODS: We distributed an online questionnaire among Latin-American pediatric surgeons. RESULTS: One hundred forty-nine pediatric surgeons from 15 countries completed the anonymous survey (71.4% of Latin-American countries), grouped into 81.9% pediatric surgeons; 8.9% pediatric colorectal surgeons, 8.9% trainees, and 1 pediatric colorectal surgeon fellow. 50.4% reported less than 5 new patients with HD per year, 36.2% 5-10 new cases, and 13.4% more than 10. Only 14.1% of surgeons have access to perform a RSB in the diagnostic work-up of patients with suspected HD, 90% in our study perform an TEB under general anesthesia. When we ask if they could perform both procedures in babies up to 6 months, 52.3% indicate that they prefer an RSB, and for patients older than 6 months, 35.4% favor an RSB. Regarding the number of samples obtained performing an TEB, 30.9% get one biopsy, 29.5% two biopsies, and 39.6% three or more samples. Surgeons obtained the most proximal biopsy at a median of 2.3 cm (range 1-4 cm) above the pectinate line. 67.8% of surgeons prescribed antibiotic prophylaxis. Overall, 16.1% experienced complications, including rectal blood loss (n = 18), and rectal perforation (n = 3). The most frequently used staining methods for rectal biopsies are hematoxylin/eosin (87%), calretinin (56, 8%), and acetylcholinesterase (21.9%). CONCLUSIONS: In Latin-America, the accessibility for RSB is limited only 18 out of 149 surgeons have access to rectal suction tool. There is no consensus regarding sample number, site of proximal biopsy, and antibiotics use. The complications associated with the procedure seems to be less than reported with RSB. Therefore, we should standardize this common surgical practice and establish universal guidelines for rectal biopsy procedure (RBP).

Comparison between Suction Rectal Biopsy and Full-Thickness Rectal Biopsy in the Diagnosis of Hirschsprung's Disease.

AIM: The aim of this study is to compare suction rectal biopsy (SRB) with full-thickness rectal biopsy (FTRB) in suspected cases of Hirschsprung's disease (HD). MATERIALS AND METHODS: Between 2014 and 2018, we enrolled 41 consecutive children with suspected HD. We analyzed demographics, sex, age, clinical symptoms, radiological images, and biopsy reports. All the children had undergone X-ray of the abdomen and pelvis and contrast enema. All of them have undergone both SRB and FTRB, and their results were compared. RESULTS: Out of 41 children, 26 were male and 15 were female. The children were aged from 5 days to 12 years. All of them presented with delayed passage of meconium, abdominal distension, and severe constipation. They all were on oral laxatives. The sensitivity and specificity of SRB are 80.95% and 90.00% when compared to FTRB which has 100% and 100%, respectively. CONCLUSION: FTRB is the gold standard test for diagnosing HD. SRB may be a good screening test in suspected HD cases. SRB is not as equal and effective as FTRB for diagnosing HD.

Diagnosis of Hirschsprung Disease.

Diagnosis or exclusion of Hirschsprung disease (HSCR) is a frequent exercise in any pediatric hospital. Although HSCR may present at different ages and with varied clinical findings, the most common presentation is a neonate with severe constipation or signs of intestinal obstruction. A variety of diagnostic tests including contrast enema and anorectal manometry may be used as diagnostic screens, but diagnosis ultimately rests upon histopathological evaluation of a rectal biopsy. For the experienced pathologist, conventional hematoxylin-and-eosin-stained sections often suffice to exclude HSCR or establish the diagnosis. However, ancillary diagnostic tests such as acetylcholinesterase histochemistry or calretinin immunohistochemistry are complementary and extremely helpful in some cases. In this Perspectives article, we review the clinical and pathological features of HSCR, highlight those that are found in most patients, and discuss how to address particularly challenging aspects of the diagnostic workup.

A comparison of suction and full-thickness rectal biopsy in children.

BACKGROUND: Rectal biopsy evaluation by an experienced pathologist is the gold standard in diagnosis of Hirschsprung's disease (HD). Although both suction rectal biopsy (SRB) and full-thickness (FTRB) rectal biopsy are performed, the ability for SRB to obtain adequate tissue in older children has been questioned. We hypothesized that SRB and FTRB yield tissue specimens of different size but are equally adequate for diagnosis. METHODS: Records of children who underwent rectal biopsy to evaluate for HD between January 2007 and July 2014 were reviewed. Volume, percent submucosa, and specimen adequacy were compared between biopsy techniques, and the effect of age on biopsy adequacy was assessed. Data were analyzed by mixed-effects models with covariate adjustment for age at biopsy and Fisher's exact test. RESULTS: Forty-seven children underwent a total of 58 biopsies, 45 SRB and 13 FTRB. Thirty-seven were performed before 12 mo of age, and 21 after 12 mo of age. Volume of SRB specimens was significantly smaller than FTRB across ages (14.8 ± 7.8 mm(3)versus 121.3 ± 13.8 mm(3), P = 0.0001). Percent submucosa did not differ significantly between SRB and FTRB specimens across ages (63.8 ± 2.7% versus 66.5 ± 4.3%, P = 0.575). The number of inadequate biopsies was low and not significantly different across ages (P = 0.345), or when comparing by biopsy method (P = 0.689). All biopsies were clinically diagnostic. There were no complications. CONCLUSIONS: Tissue specimens obtained by SRB are smaller than those obtained by FTRB, especially in older children. SRB and FTRB appear equivalent in their ability to provide adequate submucosa. Differences in cost and patient satisfaction between rectal biopsy techniques must be studied to further define the best overall technique.

Aganglionosis with the absence of hypertrophied nerve fibres predicts disease proximal to rectosigmoid colon.

PURPOSE: The gold standard for the diagnosis of Hirschsprung's disease (HSCR) is the pathologic evaluation of a rectal biopsy that demonstrates the absence of ganglion cells and nerve fibre hypertrophy. However, it has been frequently reported that hypertrophic nerves may not be present in some variants like long-segment HSCR, total colonic aganglionosis, premature and very young infants. The aim of this study was to determine this association. METHODS: We performed a retrospective review of the HSCR database at our tertiary care children's hospital from 2000 to 2013. In order to analyse the relationship between the diameter of the nerve fibres and the level of aganglionosis, we classified the patient sample into two groups-fibres ≤40 and >40 µm. The groups were statistically compared with P < 0.05 being significant. RESULTS: Rectal biopsies of 92 patients confirmed as HSCR with definitive operation performed at the same institution were reviewed. The mean nerve diameter was 50.1 µm (range 20-87.5 µm). Nerve fibre diameter ≤40 µm was predictive of transition zone above the sigmoid colon. A specificity of 77.3 % and a likelihood ratio of 2.03 supported this perception. No correlation was noted between nerve fibre diameter and gestational age at birth, birth weight or age at biopsy. CONCLUSION: The absence of nerve fibre hypertrophy in the presence of aganglionosis on rectal biopsy specimens is predictive of long-segment HSCR.

The Diagnostic Pathway of Hirschsprung's Disease in Paediatric Patients: A Single-Centre Experience.

BACKGROUND: The presenting symptoms of patients with Hirschsprung's disease (HD) are a failure to pass meconium, abdominal distension, and bilious vomiting. The gold standard diagnosis is a rectal biopsy to confirm aganglionosis. The aim of our study was to describe the diagnostic pathway of Hirschsprung's disease at our institution and document the indication for a rectal biopsy. METHODS: We have performed a prospective collection of all patients who underwent a rectal biopsy to exclude HD from December 2022 until September 2023 including. The following data were collected: patient's age, presenting symptoms, type of biopsy, failure rate, complications, and histopathological results. RESULTS: We identified 33 patients who underwent 34 rectal biopsies at 0.6 years of age. A total of 17 patients had a rectal suction biopsy (RSB), and 17 patients underwent a partial thickness under general anaesthesia (GA). 1/17 (6%) patients had an inconclusive RSB and subsequently underwent a biopsy under GA. Constipation and chronic abdominal distension plus vomiting were the most common presenting symptoms throughout all ages. Five patients (15%) had a rectal biopsy that was positive for HD. CONCLUSION: A protocolised approach to the assessment of infants and children with suspected HD ensures the appropriate utilisation of invasive procedures such as biopsy.

Diagnostic accuracy of barium enema versus full-thickness rectal biopsy in children with clinically suspected Hirschsprung's disease: A comparative cross-sectional study.

Background and Aims: Hirschsprung's disease (HSD) remains a common cause of pediatric intestinal obstruction. Barium contrast enema (BE) is the primary imaging modality for the evaluation of clinically suspected cases. Here, we aimed to assess the diagnostic accuracy of BE in children with clinically suspected HSD when compared to a gold standard full-thickness rectal biopsy (FTRB). Methods: We recruited and consecutively enrolled children with clinically suspected HSD at two tertiary teaching hospitals. Participants underwent BE imaging and two radiologists interpreted the findings independently. Participants further underwent FTRB by pediatric surgeons as the confirmatory test. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and receiver operating characteristics (ROC) with the area under the curve (AUC) were calculated on Stata version 14.2, taking FTRB as the standard. Results: We enrolled 55 cases, of which 49 completed the evaluation and were included in the final analysis. The median age was 9.4 months (interquartile range: 2-24], with a male-to-female ratio of 4.4:1. The sensitivity, specificity, PPV, and NPV of BE were 0.95 (95% confidence interval [CI] [0.81-0.99]), 0.73 (95% CI [0.39-0.94]), 0.92 (95% CI [0.82-0.97]), and 0.80 (95% CI [0.50-0.94]), respectively. On AUC, the diagnostic accuracy of BE compared to the confirmatory FTRB was 0.84 (95% CI [0.69-0.98]). The diagnostic accuracy was higher in neonates (ROC: 1.00) when compared to infants (ROC: 0.83) or those above 1 year of age (ROC: 0.798). HSD-suggestive BE findings were associated with absence of ganglion cells on FTRB (χ 2 = 23.301, p < 0.001). Inverted rectosigmoid ratio and transition zone were more sensitive in detecting HSD of 0.92 (95% CI [0.74-0.98]) and 0.81 (95% CI [0.63-0.92]), respectively. Conclusion: BE is sufficiently accurate in the diagnosis of children with HSD, suggesting BE would likely be used to inform surgical management in settings where confirmatory biopsy is lacking. However, clinical judgment is warranted in interpreting negative BE findings.

Early Detection and Intervention for Hirschsprung's Disease: A Key to Successful Outcomes.

Hirschsprung's disease is a congenital disorder characterized by the absence of nerve cells in the colon muscles, leading to difficulties in evacuating stool. This case report describes a newborn patient presenting with typical symptoms of the disease, including abdominal distension, constipation, diarrhea, and fever. The patient's initial laboratory investigations revealed anemia (Hb: 9.80 g/dL), low RBC count (3.50 million/cu mm), elevated RDW (16.70%), increased WBC count (11 000/cu mm), and raised platelet levels (891 000/µL) along with an elevated CRP (3.22). The baby received a comprehensive treatment regimen, including blood transfusion, Syp. MVBC, Enterogermina, Inj. Pan, Inj. Metro, Inj. Piptaz, Inj. Vancomycin, Calcium gluconate, and Inj. Aminoven. Following this treatment and necessary surgical intervention, the patient demonstrated significant improvement in frequent bowel movements and alleviation of other symptoms. This case highlights the importance of prompt diagnosis and multidisciplinary management for favorable outcomes in infants with Hirschsprung's disease.

Centralized intestinal organoid generation is a feasible and safe approach for personalized medicine as demonstrated in the HIT-CF Europe Organoid Study.

BACKGROUND: Patient-derived intestinal organoids (PDIOs) show great potential as in vitro drug testing platform for personalised medicine in Cystic Fibrosis and oncology. PDIOs can be generated by culturing adult stem cells obtained through rectal forceps biopsy or suction biopsy, but the safety of these procedures and the success rates of generating organoids after shipment to a centralized lab using these procedures has not been studied in this context. We here report the safety and success rates of both biopsy procedures and the subsequent generation of PDIOs in the international multicentre HIT-CF Organoid Study. METHODS: 502 biopsy procedures were conducted, on 489 adult people with Cystic Fibrosis from 33 different hospitals across 12 countries. Depending on the preference of the hospital, either rectal forceps biopsies or suction biopsies were obtained and internationally shipped to a central laboratory for organoid generation. RESULTS: No adverse events were reported for 280 forceps biopsy procedures, while 222 rectal suction biopsy procedures resulted in 2 adverse events, namely continued bleeding and a probably nonrelated gastroenteritis. The success rate of organoid generation from all biopsies was 95%, and the main reason for failure was insufficient sample viability (3.2%). CONCLUSION: Our results indicate that both rectal suction biopsy and forceps biopsy procedures are safe procedures. The high success rates of PDIO generation from the obtained tissue samples demonstrate the feasibility of the organoid technology for personalised in vitro testing in an international setting.

A current view of mitochondria damage and the diversity of lipopigment inclusions in neuronal ceroid lipofuscinose type 2 from rectal biopsy.

Neuronal ceroid lipofuscinoses (NCLs) are a growing group of neurodegenerative storage diseases, in which specific features are sought to facilitate the creation of a universal diagnostic algorithm in the future. In our ultrastructural studies, the group of NCLs was represented by the CLN2 disease caused by a defect in the TPP1 gene encoding the enzyme tripeptidyl-peptidase 1. A 3.5-year-old girl was affected by this disease. Due to diagnostic difficulties, the spectrum of clinical, enzymatic, and genetic tests was extended to include analysis of the ultrastructure of cells from a rectal biopsy. The aim of our research was to search for pathognomonic features of CLN2 and to analyse the mitochondrial damage accompanying the disease. In the examined cells of the rectal mucosa, as expected, filamentous deposits of the curvilinear profile (CVP) type were found, which dominated quantitatively. Mixed deposits of the CVP/fingerprint profile (FPP) type were observed less frequently in the examined cells. A form of inclusions of unknown origin, not described so far in CLN2 disease, were wads of osmophilic material (WOMs). They occurred alone or co-formed mixed deposits. In addition, atypically damaged mitochondria were observed in muscularis mucosae. Their deformed cristae had contact with inclusions that looked like CVPs. Considering the confirmed role of the c subunit of the mitochondrial ATP synthase in the formation of filamentous lipopigment deposits in the group of NCLs, we suggest the possible significance of other mitochondrial proteins, such as mitochondrial contact site and cristae organizing system (MICOS), in the formation of these deposits. The presence of WOMs in the context of searching for ultrastructural pathognomonic features in CLN2 disease also requires further research.

The prevalence of Hirschsprung disease in premature infants after suction rectal biopsy.

BACKGROUND: The prevalence of Hirschsprung disease (HD) in the premature infant population is not well documented. However, delayed passage of stool is common in premature infants, and suction rectal biopsy (SRB) is often used to evaluate for HD in this population. The use of SRB is unknown. Therefore, we evaluated the role of SRB in premature infants with abnormal stooling patterns. METHODS: After Institutional Review Board approval, a retrospective study was conducted on all infants having an SRB performed to exclude HD from January 2000 to December 2010. Infants were divided into two groups according to gestational age (premature < 37 wk; term ≥ 37 wk). Demographics, diagnosis, treatments, and outcomes were collected. A subset analysis was performed on patients diagnosed with HD. RESULTS: Two hundred sixty-nine infants were identified (113 premature and 156 term). Six premature infants (5.3%) and 79 term infants (50.6%) were found to have HD (P < 0.01). As expected, gestational age was significantly different between groups (31.7 versus 38.9 wk, P < 0.01) (Table 1). Premature infants were less likely to have prenatal care (35% versus 55%, P < 0.01) and had longer lengths of hospital stay (45.6 versus 17.6 d, P < 0.01). The most common location of aganglionosis was rectosigmoid in both groups (group 1, 50%; group 2, 33%, P = 0.7). CONCLUSIONS: HD occurs significantly less often in premature infants than in term infants. Alternative diagnoses should be investigated in this population when delayed stooling patterns are encountered. SRB should be used more selectively in this group.