Development and application of one-step multiplex Real-Time PCR for detection of three main pathogens associated with bovine neonatal diarrhea.

Introduction: Neonatal calf diarrhea (NCD) is one of the most common diseases in calves, causing huge economic and productivity losses to the bovine industry worldwide. The main pathogens include bovine rotavirus (BRV), bovine coronavirus (BCoV), and Enterotoxigenic Escherichia coli (ETEC) K99. Since multiple infectious agents can be involved in calf diarrhea, detecting each causative agent by traditional methods is laborious and expensive. Methods: In this study, we developed a one-step multiplex Real-Time PCR assay to simultaneously detect BRV, BCoV, and E. coli K99+. The assay performance on field samples was evaluated on 1100 rectal swabs of diseased cattle with diarrhea symptoms and compared with the conventional gel-based RT-PCR assay detect BRV, BCoV, and E. coli K99+. Results: The established assay could specifically detect the target pathogens without cross-reactivity with other pathogens. A single real-time PCR can detect ~1 copy/µL for each pathogen, and multiplex real-time PCR has a detection limit of 10 copies/µL. Reproducibility as measured by standard deviation and coefficient of variation were desirable. The triple real-time PCR method established in this study was compared with gel-based PT-PCR. Both methods are reasonably consistent, while the real-time PCR assay was more sensitive and could rapidly distinguish these three pathogens in one tube. Analysis of surveillance data showed that BRV and BCoV are major enteric viral pathogens accounting for calves' diarrhea in China. Discussion: The established assay has excellent specificity and sensitivity and was suitable for clinical application. The robustness and high-throughput performance of the developed assay make it a powerful tool in diagnostic applications and calf diarrhea research. ​.

The Use of Diets in the Diagnosis and Treatment of Common Gastrointestinal Diseases in Dogs and Cats.

The nutritional health of dogs and cats is important to pet owners around the world. Nutrition is inextricably linked to the health of the gastrointestinal system and vice versa. Gastrointestinal signs, such as vomiting, diarrhea, anorexia, or weight loss, are one of the most common reasons that dog and cat owners make non-routine appointments with veterinarians. Those patients are evaluated systematically to identify and/or rule out the causes of the symptoms. Some causes of chronic diarrhea are within the gastrointestinal tract while others are secondary to pathogenic factors outside the digestive system. Some useful biomarkers of chronic intestinal disease (enteropathy) exist in serum and feces. After determination that the clinical signs are due to primary gastrointestinal disease and that there is no parasitism, specific diets are used for at least two weeks. There are several types of diets for pets with chronic enteropathies. There are limited ingredient diets and hydrolyzed protein diets with reduced levels of allergens. There are also highly digestible and fiber-enhanced diets. Some diets contain probiotics and/or prebiotics. If symptoms do not improve and the patient is stable, a diet from a different class may be tried. For chronic enteropathies, the prognosis is generally good for symptom resolution or at least improvement. However, if interventions with novel diets do not ameliorate the symptoms of chronic enteropathy, then antibiotic, anti-inflammatory, or immunosuppressant therapy or further, more invasive diagnostics such as taking an intestinal biopsy, may be indicated. Pancreatitis is a common gastrointestinal disease in dogs and cats and patients may present with mild to severe disease. Many patients with mild to moderate disease can be successfully treated with early supportive care, including feeding a low-fat diet. A novel pharmaceutical, fuzapladib (Panoquell-CA1) looks very promising for treating more severe forms of acute pancreatitis in dogs. Maintenance on a low-fat diet may prevent pancreatitis in at-risk dogs. Future advances in medicine will allow pet owners and veterinarians to use dietary management to maximize the health of their dogs and cats.

Pyrococcus furiosus Argonaute Based Detection Assays for Porcine Deltacoronavirus.

Porcine deltacoronavirus (PDCoV) is a major cause of diarrhea and diarrhea-related deaths among piglets and results in massive losses to the overall porcine industry. The clinical manifestations of porcine diarrhea brought on by the porcine epidemic diarrhea virus (PEDV), porcine transmissible gastroenteritis virus (TGEV), and PDCoV are oddly similar to each other. Hence, the identification of different pathogens through molecular diagnosis and serological techniques is crucial. Three novel detection methods for identifying PDCoV have been developed utilizing recombinase-aided amplification (RAA) or reverse transcription recombinase-aided amplification (RT-RAA) in conjunction with Pyrococcus furiosus Argonaute (PfAgo): RAA-PfAgo, one-pot RT-RAA-PfAgo, and one-pot RT-RAA-PfAgo-LFD. The indicated approaches have a detection limit of around 60 copies/µL of PDCoV and do not cross-react with other viruses including PEDV, TGEV, RVA, PRV, PCV2, or PCV3. The applicability of one-pot RT-RAA-PfAgo and one-pot RT-RAA-PfAgo-LFD were examined using clinical samples and showed a positive rate comparable to the qPCR method. These techniques offer cutting-edge technical assistance for identifying, stopping, and managing PDCoV.

Field evaluation of a simple and rapid diagnostic test, RLDT to detect Shigella and enterotoxigenic E. coli in Indian children.

The diagnostic assays currently used to detect Shigella spp. (Shigella) and enterotoxigenic Escherichia coli (ETEC) are complex or elaborate which make them difficult to apply in resource poor settings where these diseases are endemic. The simple and rapid nucleic acid amplification-based assay "Rapid LAMP-based Diagnostic Test (RLDT)" was evaluated to detect Shigella spp (Shigella) and enterotoxigenic Escherichia coli (ETEC) and determine the epidemiology of these pathogens in Kolkata, India. Stool samples (n = 405) from children under five years old with diarrhea seeking care at the hospitals were tested, and 85(21%) and 68(17%) by RLDT, 91(23%) and 58(14%) by quantitative PCR (qPCR) and 35(9%) and 15(4%) by culture, were positive for Shigella and ETEC, respectively. The RLDT showed almost perfect agreement with qPCR, Kappa 0.96 and 0.89; sensitivity 93% and 98%; specificity 100% and 97% for Shigella and ETEC, respectively. While RLDT detected additional 12% Shigella and 13% ETEC than culture, all culture positives for Shigella and ETEC except one each were also positive by the RLDT, sensitivity 97% and 93% respectively. RLDT is a simple, sensitive, and rapid assay that could be implemented with minimum training in the endemic regions to strengthen the disease surveillance system and rapid outbreak detection.

Prolonged Intake of Luvos Healing Earth does not alter the Composition of the Gut Microbiota in Patients with Diarrhea-predominant Irritable Bowel Syndrome and Healthy Controls.

BACKGROUND AND AIMS: The mineral compound Luvos Healing Earth (LHE) is a commercially available remedy empirically used for a variety of gastrointestinal disorders. The aim of this study was to investigate the possible effect of prolonged LHE therapy on gut microbiota in healthy individuals and in patients with diarrhea-predominant irritable bowel syndrome (IBS-D). METHODS: In this prospective exploratory study, a total of 20 participants, including 12 healthy controls and 8 patients with IBS-D, received treatment with LHE (Magenfein Granulat, 1 sachet bid) for 6 weeks. Fecal samples were collected for microbiota analysis in the morning fasting state at regular intervals at 6 different timepoints: 2 weeks before starting therapy (Screen), and every 2 weeks during LHE therapy (V0-V3). Additionally, a follow-up visit was scheduled 4 weeks after the end of treatment (V4). Microbiota analysis was performed using the GA-map® Dysbiosis Test Lx v2. Dysbiosis Index, bacterial diversity, as well as the balance or imbalance of functionally important bacteria were assessed. RESULTS: The microbiota analysis revealed an overlap in gut microbiota profiles between healthy controls and patients with IBS-D. Bacterial communities were consistently stable during the entire treatment period, and no significant variations in composition were observed 4 weeks after the end of the therapeutic intervention. There was a remarkable stability of microbiota profiles over time within each individual and a high inter-individual variation. The majority of fecal samples exhibited profiles, reflecting an eubiotic state, with no significant changes in dysbiosis index, functional bacteria profiles, or bacterial diversity. CONCLUSION: Our findings indicate intraindividual resilience of microbiota consortia during the entire study period. Prolonged intake of LHE does not cause significant alterations in fecal microbiota profiles in healthy controls and patients with IBS-D. Luvos Healing Earth does not affect the stability of gut microbial diversity and bacterial functions.

Gender differences in gastrointestinal, biopsychosocial and healthcare-seeking behaviors in Chinese patients with irritable bowel syndrome predominant with diarrhea.

BACKGROUND: Evidences of comparison of sex difference in Chinese irritable bowel syndrome (IBS) patients were few. We aim to compare gender difference in the biopsychosocial characteristics of Chinese patients of IBS predominant with diarrhea (IBS-D). METHODS: IBS-D patients meeting Rome III criteria were enrolled. We administered IBS symptom questionnaires, evaluation of psychological status (HAMD and HAMA scales) and IBS quality of life (IBS-QOL), dietary habits, healthcare seeking behaviors, and compared biopsychosocial characteristics between male and female patients. RESULTS: Four hundred and ninety patients were enrolled including 299 males and 191 females. More female patients reported abdominal pain associated with defecation (84.3% vs. 74.9%, P = 0.014) while males reported more abdominal discomfort (39.8% vs. 26.7%, P = 0.003). Females had higher IBS symptom score (9.7 ± 1.7 vs. 9.4 ± 1.4, P = 0.025) and more of females had severe abdominal pain/discomfort (17.8% vs. 12.4%, P = 0.013) while there were no significant differences of other bowel symptoms. Females reported higher incidence of comorbid anxiety state (64.9% vs. 52.8%, P = 0.008) and depression state (35.6% vs. 19.7%, P < 0.001) than males. Female patients also had lower IBS-QOL score (70.2 ± 20.4 vs. 75.1 ± 16.8, P = 0.028) and more frequent consultations, as well as less response for dietary modification than males. CONCLUSIONS: Chinese female patients with IBS-D had more prominent psychosocial disorders compared to male patients and their abdominal symptoms had minor differences.

[Chronic diarrhoea: when is it celiac disease and when not?] / Chronische Durchfälle: Wann ist es eine Zöliakie und wann nicht?

Patients who come to clinical consultation for chronic diarrhoea (i.e., diarrhoea lasting for more than four weeks) may suffer from a wide range of clinical conditions. The possible diagnoses range from a misunderstanding of what can be considered normal and what pathological in terms of daily bowel movements, to a severe malabsorption syndrome. Since the list of possible causes of chronic diarrhoea can be puzzling, the physician's approach needs to be systematic and structured in order to allow the correct diagnosis and treatment. This article proposes an algorithm for the diagnosis of chronic diarrhoea and discusses in detail the key clinical aspects of celiac disease, which is considered a paradigmatic disease as regards chronic malabsorptive diarrhoea.

Defining a metagenomic threshold for detecting low abundances of Providencia alcalifaciens in canine faecal samples.

Introduction: Acute haemorrhagic diarrhoea syndrome (AHDS) in dogs is a condition of unknown aetiology. Providencia alcalifaciens is suspected to play a role in the disease as it was commonly found in dogs suffering from AHDS during a Norwegian outbreak in 2019. The role of this bacterium as a constituent of the canine gut microbiota is unknown, hence this study set out to investigate its occurrence in healthy dogs using metagenomics. Materials and methods: To decrease the likelihood of false detection, we established a metagenomic threshold for P. alcalifaciens by spiking culture-negative stool samples with a range of bacterial dilutions and analysing these by qPCR and shotgun metagenomics. The detection limit for P. alcalifaciens was determined and used to establish a metagenomic threshold. The threshold was validated on naturally contaminated faecal samples with known cultivation status for P. alcalifaciens. Finally, the metagenomic threshold was used to determine the occurrence of P. alcalifaciens in shotgun metagenomic datasets from canine faecal samples (n=362) collected in the HUNT One Health project. Results: The metagenomic assay and qPCR had a detection limit of 1.1x103 CFU P. alcalifaciens per faecal sample, which corresponded to a Cq value of 31.4 and 569 unique k-mer counts by shotgun metagenomics. Applying this metagenomic threshold to 362 faecal metagenomic datasets from healthy dogs, P. alcalifaciens was found in only 1.1% (95% CI [0.0, 6.8]) of the samples, and then in low relative abundances (median: 0.04%; range: 0.00 to 0.81%). The sensitivity of the qPCR and shotgun metagenomics assay was low, as only 40% of culture-positive samples were also positive by qPCR and metagenomics. Discussion: Using our detection limit, the occurrence of P. alcalifaciens in faecal samples from healthy dogs was low. Given the low sensitivity of the metagenomic assay, these results do not rule out a significantly higher occurrence of this bacterium at a lower abundance.

Development and Evaluation of a Rapid GII Norovirus Detection Method Based on CRISPR-Cas12a.

Norovirus is highly infectious and rapidly transmissible and represents a major pathogen of sporadic cases and outbreaks of acute gastroenteritis worldwide, causing a substantial disease burden. Recent years have witnessed a dramatic increase in norovirus outbreaks in China, significantly higher than in previous years, among which GII norovirus is the predominant prevalent strain. Therefore, rapid norovirus diagnosis is critical for clinical treatment and transmission control. Hence, we developed a molecular assay based on RPA combined with the CRISPER-CAS12a technique targeting the conserved region of the GII norovirus genome, the results of which could be displayed by fluorescence curves and immunochromatographic lateral-flow test strips. The reaction only required approximately 50 min, and the results were visible by the naked eye with a sensitivity reaching 102 copies/µl. Also, our method does not cross-react with other common pathogens that cause intestinal diarrhea. Furthermore, this assay was easy to perform and inexpensive, which could be widely applied for detecting norovirus in settings including medical institutions at all levels, particularly township health centers in low-resource areas.

Comparison of metagenomic and traditional methods for diagnosis of E. coli enteric infections.

Diarrheagenic Escherichia coli, collectively known as DEC, is a leading cause of diarrhea, particularly in children in low- and middle-income countries. Diagnosing infections caused by different DEC pathotypes traditionally relies on the cultivation and identification of virulence genes, a resource-intensive and error-prone process. Here, we compared culture-based DEC identification with shotgun metagenomic sequencing of whole stool using 35 randomly drawn samples from a cohort of diarrhea-afflicted patients. Metagenomic sequencing detected the cultured isolates in 97% of samples, revealing, overall, reliable detection by this approach. Genome binning yielded high-quality E. coli metagenome-assembled genomes (MAGs) for 13 samples, and we observed that the MAG did not carry the diagnostic DEC virulence genes of the corresponding isolate in 60% of these samples. Specifically, two distinct scenarios were observed: diffusely adherent E. coli (DAEC) isolates without corresponding DAEC MAGs appeared to be relatively rare members of the microbiome, which was further corroborated by quantitative PCR (qPCR), and thus unlikely to represent the etiological agent in 3 of the 13 samples (~23%). In contrast, ETEC virulence genes were located on plasmids and largely escaped binning in associated MAGs despite being prevalent in the sample (5/13 samples or ~38%), revealing limitations of the metagenomic approach. These results provide important insights for diagnosing DEC infections and demonstrate how metagenomic methods can complement isolation efforts and PCR for pathogen identification and population abundance. IMPORTANCE: Diagnosing enteric infections based on traditional methods involving isolation and PCR can be erroneous due to isolation and other biases, e.g., the most abundant pathogen may not be recovered on isolation media. By employing shotgun metagenomics together with traditional methods on the same stool samples, we show that mixed infections caused by multiple pathogens are much more frequent than traditional methods indicate in the case of acute diarrhea. Further, in at least 8.5% of the total samples examined, the metagenomic approach reliably identified a different pathogen than the traditional approach. Therefore, our results provide a methodology to complement existing methods for enteric infection diagnostics with cutting-edge, culture-independent metagenomic techniques, and highlight the strengths and limitations of each approach.

A multiplex real-time RT-qPCR assay for simultaneous detection of porcine epidemic diarrhea virus, porcine deltacoronavirus, and swine acute diarrhea syndrome coronavirus.

Porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and swine acute diarrhea syndrome coronavirus (SADS-CoV) cause intestinal diseases with similar manifestations in suckling piglets. In this study, we developed a multiplex real-time PCR for differential diagnosis of PEDV, PDCoV, and SADS-CoV. The assay demonstrated high specificity with a detection limit of 5 copies/µl for each virus. The assay specifically detected PEDV, PDCoV, and SADS-CoV and excluded all other swine pathogens circulating in pigs. Furthermore, the assay exhibited satisfactory performance in analyzing clinical samples. The data indicate that the newly developed multiplex real-time PCR method can be applied for differential diagnosis of porcine enteric coronaviruses.

An isothermal nucleic acid amplification-based enzymatic recombinase amplification method for dual detection of porcine epidemic diarrhea virus and porcine rotavirus A.

Viral diarrhea is the predominant digestive tract sickness in piglings, resulting in substantial profit losses in the porcine industry. Porcine rotavirus A (PoRVA) and porcine epidemic diarrhea virus (PEDV) are the main causes of grave gastroenteritis and massive dysentery, especially in piglets. PoRVA and PEDV have high transmissibility, exhibit similar clinical symptoms, and frequently co-occur. Therefore, to avoid financial losses, a quick, highly efficient, objective diagnostic test for the prevention and detection of these diseases is required. Enzymatic recombinase amplification (ERA) is a novel technology based on isothermal nucleic acid amplification. It demonstrates high sensitivity and excellent specificity, with a short processing time and easy operability, compared with other in vitro nucleic acid amplification technologies. In this study, a dual ERA method to detect and distinguish between PEDV and PoRVA nucleic acids was established. The method shows high sensitivity, as the detection limits were 101 copies/µL for both viruses. To test the usefulness of this method in clinical settings, we tested 64 swine clinical samples. Our results were 100% matched with those acquired using a commercially available kit. Therefore, we have successfully developed a dual diagnostic ERA nucleic acids method for detecting and distinguishing between PEDV and PoRVA.

Utility of irritable bowel syndrome subtypes and most troublesome symptom in predicting disease impact and burden.

BACKGROUND: Little is known about the characteristics of individuals with irritable bowel syndrome (IBS) according to stool subtype or the most troublesome symptom reported by the individual, or whether these are useful in predicting the impact of IBS. METHODS: We collected demographic, gastrointestinal, and psychological symptoms, healthcare usage and direct healthcare costs, impact on work and activities of daily living, and quality of life data from individuals with Rome IV-defined IBS. KEY RESULTS: We recruited 752 people with Rome IV IBS. Individuals with IBS-D reported a poorer disease-specific quality of life than those with IBS-C or IBS-M (mean (SD) IBS-QOL 45.3 (23.0) for IBS-D, vs. 52.3 (19.9) for IBS-C, vs. 49.4 (22.0) for IBS-M, p = 0.005). Mean (SD) IBS-QOL scores were also lower amongst those who reported diarrhea (44.8 (22.3)) or urgency (44.6 (22.3)) as their most troublesome symptom, compared with those reporting abdominal pain (52.2 (22.9)), constipation (49.5 (21.8)), or abdominal bloating or distension (50.4 (21.3)). However, there were no differences in mean EQ-5D scores, IBS severity, levels of anxiety, depression, somatoform symptom-reporting, or gastrointestinal symptom-specific anxiety. Direct healthcare costs of IBS were similar across all subtypes and all most troublesome symptom groups, although some differences in work productivity and social leisure activities were detected. CONCLUSIONS AND INFERENCES: There appears to be limited variation in the characteristics of individuals with Rome IV IBS based on both stool subtypes and most troublesome symptom reported, suggesting that gastrointestinal symptoms alone have limited ability to predict disease impact and burden.

Evaluation of a novel triplex immunochromatographic test for rapid simultaneous detection of norovirus, rotavirus, and adenovirus on a single strip test.

BACKGROUND: Acute gastroenteritis is one of the major causes of morbidity and mortality in young children worldwide. Among these, rotavirus, norovirus, and adenovirus have been reported as the primary viral pathogens associated with the disease. Rapid diagnosis of viral pathogens is crucial when diarrhea outbreaks occur to ensure the timely administration of appropriate treatment and control measures. METHODS: We evaluated three immunochromatographic test kits designed for the detection of norovirus, rotavirus, and adenovirus in 71 stool specimens collected from children with diarrhea who visited clinics in Japan. The first kit is a triplex immunochromatographic test kit designed for simultaneous detections of norovirus, rotavirus, and adenovirus on a single strip (this kit was referred to as IC-A). The other two immunochromatographic test kits are a dual detection kit for rotavirus and adenovirus, and a single detection kit for norovirus (IC-B). The RT-PCR/PCR was used as the gold standard method. RESULTS: The results revealed that both IC-A and IC-B kits exhibited the same level of sensitivity of detection for rotavirus (72.7%) and adenovirus (22.7%), although the detection rate was lower than that of the RT-PCR/PCR method. However, there was a slight difference in the sensitivity of detection for norovirus between IC-A and IC-B, at 86.7% and 93.3%, respectively. The sensitivity of detection for adenovirus of both kits was relatively lower than those of RT-PCR method. This could be due to low viral load of adenovirus in clinical specimens below the detection limit of IC-A and IC-B kits. However, both immunochromatographic test kits (IC-A and IC-B) exhibited 100% specificity for norovirus, rotavirus, and adenovirus. CONCLUSIONS: The triplex immunochromatographic test kit (IC-A) designed for simultaneous detection of norovirus, rotavirus, and adenovirus has been proved to be more practical and convenient than the use of single or dual detection kits with more or less the same sensitivity and specificity of detections.

Enhancing enteric pathogen detection: implementation and impact of multiplex PCR for improved diagnosis and surveillance.

BACKGROUND: Syndromic surveillance of acute gastroenteritis plays a significant role in the diagnosis and management of gastrointestinal infections that are responsible for a substantial number of deaths globally, especially in developing countries. In Lebanon, there is a lack of national surveillance for acute gastroenteritis, and limited data exists regarding the prevalence of pathogens causing diarrhea. The one-year study aims to investigate the epidemiology of common gastrointestinal pathogens and compare our findings with causative agents of diarrhea reported by our study collaborative centers. METHODS: A multicenter, cross-sectional study was conducted over a one-year period. A total of 271 samples were obtained from outpatients and inpatients presenting with symptoms of acute gastroenteritis at various healthcare facilities. The samples were then analyzed using Allplex gastrointestinal assay that identifies a panel of enteric pathogens. RESULTS: Overall, enteropathogens were detected in 71% of the enrolled cases, 46% of those were identified in patients as single and 54% as mixed infections. Bacteria were observed in 48%, parasites in 12% and viruses in 11%. Bacterial infections were the most prevalent in all age groups. Enteroaggregative E. coli (26.5%), Enterotoxigenic E. coli (23.2%) and Enteropathogenic E. coli (20.3%) were the most frequently identified followed by Blastocystis hominis (15.5%) and Rotavirus (7.7%). Highest hospitalization rate occurred with rotavirus (63%), Enterotoxigenic E. coli (50%), Blastocystis hominis (45%) and Enteropathogenic E. coli (43%). Enteric pathogens were prevalent during summer, fall and winter seasons. CONCLUSIONS: The adoption of multiplex real-time PCR assays in the diagnosis of gastrointestinal infections has identified gaps and improved the rates of detection for multiple pathogens. Our findings highlight the importance of conducting comprehensive surveillance to monitor enteric infections. The implementation of a syndromic testing panel can therefore provide healthcare professionals with timely and accurate information for more effective treatment and public health interventions.

Multicenter evaluation of BioCode GPP for syndromic molecular detection of gastrointestinal pathogens from stool specimens.

Acute gastroenteritis (AGE) is a leading cause of morbidity and mortality worldwide across all age groups that disproportionally affects young children in low- and middle-income countries and immunocompromised patients in high-income countries. Regional outbreaks of AGE are typically detected by traditional microbiological detection methods that target limited organisms and are associated with low sensitivity and lengthy time-to-results. Combined, these may result in repeat testing, imprecise or delayed treatment, and delayed recognition of outbreaks. We conducted a multi-site prospective study comparing the BioCode Gastrointestinal Pathogen Panel (BioCode GPP) for the detection of 17 common bacterial, viral, and protozoan causes of gastroenteritis with reference methods, including stool culture, enzyme immunoassays, pathogen-specific PCR assays, and sequencing. One thousand five hundred fifty-eight residual, de-identified stool samples (unpreserved stool and stool in Cary-Blair transport medium) were enrolled and tested for 11 bacterial, 3 viral, and 3 protozoan pathogens. BioCode GPP and reference methods were positive for 392 (25.2%) and 283 (18.2%) samples, respectively (P < 0.0001). In this study, the BioCode GPP and reference methods detected 69 and 65 specimens positive for Clostridioides difficile, 51 and 48 for enteroaggregative Escherichia coli, 33 and 27 for enterotoxigenic E. coli, 50 and 47 for norovirus GI/GII, and 30 and 22 for rotavirus A, respectively. The BioCode GPP showed good positive and negative agreements for each pathogen ranging from 89.5% to 100%, with overall sensitivity and specificity of 96.1% and 99.7%, post adjudication. The BioCode GPP detected >1 pathogens in 49 samples, representing 12.5% of the total 392 positive specimens. IMPORTANCE: This study highlights performance of a novel technology for timely and accurate detection and differentiation of 17 common bacterial, viral, and protozoan causes of gastroenteritis. Utilizing molecular tests such as the BioCode Gastrointestinal Pathogen Panel may improve the detection of gastrointestinal pathogens and provide actionable results, particularly for patient populations at most risk.

Chronic diarrhoea due to trichohepatoenteric syndrome (THES) in an infant.

An infant was admitted with suspected postinfectious malabsorption with watery diarrhoea, fever and failure to thrive. She had dehydration, acute kidney injury and metabolic acidosis, which were corrected with intravenous fluids and managed with empiric antibiotics and prophylactic antifungals. She also developed Escherichia coli sepsis, meningitis and Candida skin infections during hospitalisation, which were treated according to the culture reports. Intrauterine growth restriction, woolly hair and a broad nasal bridge with chronic refractory diarrhoea prompted genetic testing to rule out syndromic diarrhoea. Whole-exome sequencing revealed a pathogenic compound heterozygous mutation causing trichohepatoenteric syndrome. She succumbed to severe infections at 80 days of life. The condition is rare, and no established guidelines or specific treatments exist; the focus is to promote optimal growth through parenteral nutrition, elemental formula and infection control. Early suspicion and molecular genetic testing can help reduce the time to diagnosis, treatment and genetic counselling.

Bile Acid Diarrhea: From Molecular Mechanisms to Clinical Diagnosis and Treatment in the Era of Precision Medicine.

Bile acid diarrhea (BAD) is a multifaceted intestinal disorder involving intricate molecular mechanisms, including farnesoid X receptor (FXR), fibroblast growth factor receptor 4 (FGFR4), and Takeda G protein-coupled receptor 5 (TGR5). Current diagnostic methods encompass bile acid sequestrants (BAS), 48-h fecal bile acid tests, serum 7α-hydroxy-4-cholesten-3-one (C4), fibroblast growth factor 19 (FGF19) testing, and 75Selenium HomotauroCholic acid test (75SeHCAT). Treatment primarily involves BAS and FXR agonists. However, due to the limited sensitivity and specificity of current diagnostic methods, as well as suboptimal treatment efficacy and the presence of side effects, there is an urgent need to establish new diagnostic and treatment methods. While prior literature has summarized various diagnostic and treatment methods and the pathogenesis of BAD, no previous work has linked the two. This review offers a molecular perspective on the clinical diagnosis and treatment of BAD, with a focus on FXR, FGFR4, and TGR5, emphasizing the potential for identifying additional molecular mechanisms as treatment targets and bridging the gap between diagnostic and treatment methods and molecular mechanisms for a novel approach to the clinical management of BAD.

Assessment of pre-referral treatment for malaria, diarrhea, and pneumonia by rural community health workers in Southwestern Uganda: a cross-sectional study.

BACKGROUND: Pre-referral treatment aims to stabilize the child's condition before transferring them to a higher level of healthcare. This study explored pre-referral treatment for diarrhea, malaria and pneumonia in children U5. The study aims to assess pre-referral treatment practices among community health workers (CHWs) for children aged 2 to 59 months diagnosed with malaria, diarrhea, and pneumonia. METHODS: Conducted in 2023, this study employed a quantitative retrospective analysis of secondary data gathered from March 2014 to December 2018. Among the subjects, 171 patients received pre-referral treatment, serving as the foundation for categorical data analysis, presenting proportions and 95% confidence intervals across different categories. RESULTS: In this cohort, 90 (53%) of the 177 children U5 were male, and age distribution showed 39 (23%), 70 (41%), and 62 (36%) in the 2-11 months, 12-35 months, and 36-60 months categories, respectively. Rapid Diagnostic Test (RDT) malaria results indicated a negative outcome in 83(60%) and positive in 55 (40%) of cases. Symptomatically, 45 (26%) had diarrhea, 52 (30%) exhibited fast breathing, and 109 (63%) presented with fever. Furthermore, 59 (35%) displayed danger signs, while 104 (61%) sought medical attention within 24 h. CONCLUSION: The study analyzed a sample of 171 children under 5 years old to assess various characteristics and variables related to pre-referral treatment. The findings reveal notable proportions in gender distribution, age categories, RDT results, presence of diarrhea, fast breathing, fever, danger signs, and timely medical visits. The results highlight the need to strengthen pre-referral treatment interventions and enhance iCCM programs.