Optical detection using CRISPR-Cas12a of Helicobacter pylori for veterinary applications.

Helicobacter pylori (H. pylori) is a zoonotic gastric microorganism capable of efficient interspecies transmission. Domesticated companion animals, particularly dogs and cats, serve as natural reservoirs for H. pylori. This phenomenon facilitates the extensive dissemination of H. pylori among households with pets. Hence, the prompt and precise identification of H. pylori in companion animals holds paramount importance for the well-being of both animals and their owners. With the assistance of Multienzyme Isothermal Rapid Amplification (MIRA) and CRISPR-Cas12a system, we successfully crafted a highly adaptable optical detection platform for H. pylori. Three sensor systems with corresponding visual interpretations were proposed. This study demonstrated a rapid turnaround time of approximately 45 min from DNA extraction to the result display. Moreover, this platform topped germiculture and real-time PCR in terms of sensitivity or efficiency in clinical diagnoses of 66 samples. This platform possesses significant potential as a versatile approach and represents the premiere application of CRISPR for the non-invasive detection of H. pylori in companion animals, thereby mitigating the dissemination of H. pylori among household members.

Relationship between gut microbiota and lung function decline in patients with chronic obstructive pulmonary disease: a 1-year follow-up study.

OBJECTIVE: Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory lung disease characterized by a persistent limitation in airflow. Gut microbiota is closely correlated with lung inflammation. However, gut microbiota has not been studied in patients with declining lung function, due to chronic lung disease progression. SUBJECTS AND METHODS: Stool samples were obtained from 55 patients with COPD that were in stable condition at enrolment (stage 1) and at a 1-year follow-up (stage 2). After extracting stool DNA, we performed next generation sequencing to analyse the distribution of gut microbiota. RESULTS: Patients were divided to control and declining lung function groups, based on whether the rate of forced expiratory volume in 1 s (FEV1) had declined over time. An alpha diversity analysis of initial and follow-up stool samples showed a significant difference in the community richness of microbiota in the declining function group, but not in the control group. At the phylum level, Bacteroidetes was more abundant in the control group and Firmicutes was more abundant in the declining function group. The Alloprevotella genus was more abundant in the control group than in the declining function group. At 1-year follow-up, the mean proportions of Acinetobacter and Stenotrophomonas significantly increased in the control and declining function groups, respectively. CONCLUSION: Some community shifts in gut microbiota were associated with lung function decline in COPD patients under regular treatment. Future studies should investigate the mechanism underlying alterations in lung function, due to changes in gut bacterial communities, in COPD.

Cryopreservation of stool samples altered the microbial viability quantitively and compositionally.

Stool is the most commonly used sample for gut microbiota analysis in humans and animals. Cryopreservation of stool at - 80 °C is a feasible and simple method in clinics and researches, especially in large-scale cohort studies. However, the viability of bacteria in stool after freezing has yet well-demonstrated quantitatively and compositionally. This study determined the viable microbiota of samples under cryopreservation at - 80 °C, relative to fresh samples and that stored at ambient. Stool samples were collected from three healthy adults. Propidium monoazide treatment combined with quantitative PCR and 16S rRNA gene sequencing was performed to target viable microbiota. After freezing, the number of viable bacteria decreased, though inter-individual difference existed. Notably, the alpha diversity of viable microbiota after freezing did not change significantly, while its composition changed. Freezing significantly reduced the viable bacteria in Gram-negative genera of Bacteroidetes and Firmicutes, and proportionally increased Gram-positive bacteria in genera of Actinobacteria and Firmicutes, including Bifidobacterium, Collinsella and Blautia, implying that the cell envelope structure associated with the bacterial sensitivity to freezing. On the contrary, the room temperature storage not only decreased the number of viable bacteria, but also decreased the microbial alpha diversity, and remarkably enriched facultative anaerobes of Escherichia-Shigella, Enterococcus and Lactococcus, some of which are opportunistic pathogens. Our findings suggested that changes in viable microbiota in stool samples caused by cryopreservation should be paid enough attention for downstream utilization.

Simple and portable on-site system for nucleic acid-based detection of Clostridium difficile in stool samples using two columns containing microbeads and loop-mediated isothermal amplification.

It is challenging to employ nucleic acid-based diagnostics for the in situ detection of Clostridium difficile from complex fecal samples because essential sample preparation and amplification procedures require various experimental resources. In this study, a simple and effective on-site nucleic acid-based detection system was used to detect C. difficile in stool samples. Two columns containing different microbeads, namely, glass and functionalized graphene oxide-coated microbeads, were designed to remove relatively large impurities by filtration and concentrate bacteria, including C. difficile, from stool samples by adsorption. The bacterial nucleic acids were effectively extracted using a small bead beater. The effectiveness of enzyme inhibitors remaining in the sample was efficiently reduced by the direct buffer developed in this study. This sample preparation kit consisting of two simple columns showed better performance in real-time polymerase chain reaction (PCR) and equivalent performance in loop-mediated isothermal amplification (LAMP) than other sample preparation kits, despite 90% simplification of the process. The amplification-ready samples were introduced into two microtubes containing LAMP pre-mixtures (one each for E. coli as an external positive control and C. difficile) by a simple sample loader, which was operated using a syringe. LAMP, which indicates amplification based on color change, was performed at 65 °C in a small water bath. The limit of detection (L.O.D) and analytical sensitivity/specificity of our simple and effective kit were compared with those of a commercial kit. C. difficile in stool samples could be detected within 1 h with 103 cfu/10 mg using LAMP combined simple on-site detection kit.

Altered Gut Archaea Composition and Interaction With Bacteria Are Associated With Colorectal Cancer.

BACKGROUND & AIMS: Changes in the intestinal microbiota have been associated with development and progression of colorectal cancer (CRC). Archaea are stable components of the microbiota, but little is known about their composition or contribution to colorectal carcinogenesis. We analyzed archaea in fecal microbiomes of 2 large cohorts of patients with CRC. METHODS: We performed shotgun metagenomic analyses of fecal samples from 585 participants (184 patients with CRC, 197 patients with adenomas, and 204 healthy individuals) from discovery (165 individuals) and validation (420 individuals) cohorts. Assignment of taxonomies was performed by exact k-mer alignment against an integrated microbial reference genome database. RESULTS: Principal component analysis of archaeomes showed distinct clusters in fecal samples from patients with CRC, patients with adenomas, and control individuals (P < .001), indicating an alteration in the composition of enteric archaea during tumorigenesis. Fecal samples from patients with CRC had significant enrichment of halophilic and depletion of methanogenic archaea. The halophilic Natrinema sp. J7-2 increased progressively in samples from control individuals, to patients with adenomas, to patients with CRC. Abundances of 9 archaea species that were enriched in fecal samples from patients with CRC distinguished them from control individuals with areas under the receiver operating characteristic curve of 0.82 in the discovery cohort and 0.83 in the validation cohort. An association between archaea and bacteria diversities was observed in fecal samples from control individuals but not from patients with CRC. Archaea that were enriched in fecal samples from patients with CRC had an extensive mutual association with bacteria that were enriched in the same samples and exclusivity with bacteria that were lost from these samples. CONCLUSIONS: Archaeomes of fecal samples from patients with CRC are characterized by enrichment of halophiles and depletion of methanogens. Studies are needed to determine whether associations between specific archaea and bacteria species in samples from patients with CRC contribute to or are a response to colorectal tumorigenesis.

Study of enteric pathogens among children in the tropics and effects of prolonged storage of stool samples.

The study was performed to compare real-time PCR after nucleic acid extraction directly from stool samples as well as from samples stored and transported on Whatman papers or flocked swabs at ambient temperature in the tropics. In addition, the possible suitability for a clear determination of likely aetiological relevance of PCR-based pathogen detections based on cycle threshold (Ct) values was assessed. From 632 Tanzanian children <5 years of age with and without gastrointestinal symptoms, 466 samples were subjected to nucleic acid extraction and real-time PCR for gastrointestinal viral, bacterial and protozoan pathogens. Equal or even higher frequencies of pathogen detections from Whatman papers or flocked swabs were achieved compared with nucleic acid extraction directly from stool samples. Comparison of the Ct values showed no significant difference according to the nucleic acid extraction strategy. Also, the Ct values did not allow a decision whether a detected pathogen was associated with gastrointestinal symptoms.

Factors affecting patient adherence to publicly funded colorectal cancer screening programmes: a systematic review.

OBJECTIVES: Colorectal cancer (CRC) is the third most common cancer. Many countries in Europe have already implemented systematic screening programmes as per the recommendations by the European Union. The impact of screening is highly dependent on participation rates. The aim of the study was to identify barriers, facilitators and modifiers to participation in systematised, stool sample-based, publicly financed CRC screening programmes. STUDY DESIGN: Systematic review. METHODS: A systematic search in PubMed, Embase, MEDLINE, CINAHL, Cochrane CENTRAL, Google Scholar and PsycINFO was undertaken. We included both qualitative and quantitative studies reporting on barriers and facilitators (excluding sociodemographic variables) to participation in stool sample-based CRC screening. Barriers and facilitators to participation were summarised and analysed. RESULTS: The inclusion criteria were met in 21 studies. Reported barriers and facilitators were categorised into the following seven themes (examples): psychology (fear of cancer), religion (believing cancer is the will of God), logistics (not knowing how to conduct the test), health-related factors (mental health), knowledge and awareness (lack of knowledge about the test), role of the general practitioner (being supported in taking the test by the general practitioner), and environmental factors (knowing someone who has participated in a screening programme). Six studies reported that non-participation was not due to a negative attitude towards screening for CRC. CONCLUSION: Many barriers to screening were found. It is important to work with peoples' fear of screening. Moreover, this review suggests that it might be possible to increase participation rates, if the population-wide awareness and knowledge of potential health benefits of CRC screening are increased and proper logistical support is provided.

Multiplex-Touchdown PCR to Simultaneously Detect Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the Major Causes of Traveler's Diarrhea.

This study aimed to develop a multiplex-touchdown PCR method to simultaneously detect 3 species of protozoan parasites, i.e., Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the major causes of traveler's diarrhea and are resistant to standard antimicrobial treatments. The target genes included the Cryptosporidium oocyst wall protein for C. parvum, Glutamate dehydrogenase for G. lamblia, and 18S ribosomal RNA (18S rRNA) for C. cayetanensis. The sizes of the amplified fragments were 555, 188, and 400 bps, respectively. The multiplex-touchdown PCR protocol using a primer mixture simultaneously detected protozoa in human stools, and the amplified gene was detected in >1×103 oocysts for C. parvum, >1×104 cysts for G. lamblia, and >1 copy of the 18S rRNA gene for C. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detect C. parvum, G. lamblia, and C. cayetanenesis in stool samples.

Physician practices in requesting stool samples for patients with acute gastroenteritis, France, August 2013-July 2014.

A better understanding of physician practices in requesting stool samples for patients with acute gastroenteritis (AG) is needed to more accurately interpret laboratory-based surveillance data. A survey was conducted in General Practitioners (GPs) between August 2013 and July 2014 to estimate the proportion of stool samples requested for patients with AG and to identify factors associated with GP requests for a stool sample. National health insurance (NHI) data together with surveillance data from a French Sentinel GP network were also used to estimate the proportion of stool samples requested. This proportion was estimated at 4·3% in the GP survey and 9·1% (95% confidence interval 8·7-9·6) using NHI data. Multivariate analysis indicated that the ratio of stool samples requested was almost five times higher in patients with bloody diarrhoea and 10-20 times higher in patients with a long duration of illness before consultation. Laboratory-based surveillance data underestimates the actual burden of disease as fewer than one in 10 AG cases consulting their GP will be requested to submit a stool sample for laboratory testing. This underestimation varies by pathogen as stool samples are more frequently requested for severe illness.

Optimised human stool sample collection for multi-omic microbiota analysis.

To accurately define the role of the gut microbiota in health and disease pathogenesis, the preservation of stool sample integrity, in terms of microbial community composition and metabolic function, is critical. This presents a challenge for any studies which rely on participants self-collecting and returning stool samples as this introduces variability and uncertainty of sample storage/handling. Here, we tested the performance of three stool sample collection/preservation buffers when storing human stool samples at different temperatures (room temperature [20 °C], 4 °C and - 80 °C) for up to three days. We compared and quantified differences in 16S rRNA sequencing composition and short-chain fatty acid profiles compared against immediately snap-frozen stool. We found that the choice of preservation buffer had the largest effect on the resulting microbial community and metabolomic profiles. Collectively analysis confirmed that PSP and RNAlater buffered samples most closely recapitulated the microbial diversity profile of the original (immediately - 80 °C frozen) sample and should be prioritised for human stool microbiome studies.

Evaluation of fecal sample collection methods for feline gut microbiome profiling: fecal loop vs. litter box.

Introduction: Microbial population structures within fecal samples are vital for disease screening, diagnosis, and gut microbiome research. The two primary methods for collecting feline fecal samples are: (1) using a fecal loop, which retrieves a rectal sample using a small, looped instrument, and (2) using the litter box, which collects stool directly from the litter. Each method has its own advantages and disadvantages and is suitable for different research objectives. Methods and results: Whole-genome shotgun metagenomic sequencing were performed on the gut microbiomes of fecal samples collected using these two methods from 10 adult cats housed in the same research facility. We evaluated the influence of collection methods on feline microbiome analysis, particularly their impact on DNA extraction, metagenomic sequencing yield, microbial composition, and diversity in subsequent gut microbiome analyses. Interestingly, fecal sample collection using a fecal loop resulted in a lower yield of microbial DNA compared to the litterbox method (p = 0.004). However, there were no significant differences between the two groups in the proportion of host contamination (p = 0.106), virus contamination (p = 0.232), relative taxonomy abundance of top five phyla (Padj > 0.638), or the number of microbial genes covered (p = 0.770). Furthermore, no significant differences were observed in alpha-diversity, beta-diversity, the number of taxa identified at each taxonomic level, and the relative abundance of taxonomic units. Discussion: These two sample collection methods do not affect microbial population structures within fecal samples and collecting fecal samples directly from the litterbox within 6 hours after defecation can be considered a reliable approach for microbiome research.

Methods, quality control and specimen management in an international multicentre investigation of type 1 diabetes: TEDDY.

BACKGROUND: The vast array and quantity of longitudinal samples collected in The Environmental Determinants of Diabetes in the Young study present a series of challenges in terms of quality control procedures and data validity. To address this, pilot studies have been conducted to standardize and enhance both biospecimen collection and sample obtainment in terms of autoantibody collection, stool sample preservation, RNA, biomarker stability, metabolic biomarkers and T-cell viability. RESEARCH DESIGN AND METHODS: The Environmental Determinants of Diabetes in the Young is a multicentre, international prospective study (n = 8677) designed to identify environmental triggers of type 1 diabetes (T1D) in genetically at-risk children from ages 3 months until 15 years. The study is conducted through six primary clinical centres located in four countries. RESULTS: As of May 2012, over three million biological samples and 250 million total data points have been collected, which will be analysed to assess autoimmunity status, presence of inflammatory biomarkers, genetic factors, exposure to infectious agents, dietary biomarkers and other potentially important environmental exposures in relation to autoimmunity and progression to T1D. CONCLUSIONS: Detailed procedures were utilized to standardize both data harmonization and management when handling a large quantity of longitudinal samples obtained from multiple locations. In addition, a description of the available specimens is provided that serve as an invaluable repository for the elucidation of determinants in T1D focusing on autoantibody concordance and harmonization, transglutaminase autoantibody, inflammatory biomarkers (T-cells), genetic proficiency testing, RNA lab internal quality control testing, infectious agents (monitoring cross-contamination, virus preservation and nasal swab collection validity) and HbA1c testing.

Yield of Emergency Department Stool Culture Tests Among Children With Acute Gastroenteritis in Israel.

Previous studies have attempted to predict a positive stool culture in pediatric patients with acute gastroenteritis (AGE), but most of them are either from developing countries or are outdated. In all, 276 patients with AGE and 560 control patients were analyzed for differences in clinical factors including the presence of fever, highest recorded temperature, bloody diarrhea, number of bowel movements in 24 hours prior to presentation, and the presence of seizures, as well as laboratory parameters including leukocyte count and C-reactive protein (CRP). Positive stool sample rate was 13.7%. The most common bacterial pathogen was Campylobacter jejuni. Bacterial AGE was significantly associated with fever >37.9°C, bloody diarrhea, higher stool passing frequency, seizures, and CRP levels. For pediatric patients who present to the emergency department with AGE and present without bloody diarrhea, fever, frequent stool passing, or seizures, a stool culture test is of poor yield and may not be necessary.

Impact of COVID-19 Restrictions on Acute Gastroenteritis in Children: A Regional, Danish, Register-Based Study.

This study aimed to evaluate the impact of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) restrictions such as social distancing on the occurrence of acute gastroenteritis (AGE) among children. This study is a register-based study, including every child seen in the departments of paediatrics with the initial diagnosis of AGE in three neighbouring hospitals in Denmark, from March 2018 through February 2021. The study also included every positive stool sample for AGE-causing pathogens analysed in these three hospitals from children during the same period. The Wilcoxon rank-sum test was used to determine differences between the period during the SARS-CoV-2 restrictions and before. In all, 222,157 children were seen in the three paediatric departments during this period. Of these, 3917 children were diagnosed with AGE. We found a decrease of 46.6% in AGE-related visits per month after the SARS-CoV-2 restrictions were introduced compared to before (p-value < 0.001). Positive stool samples decreased by 38.2% (p-value = 0.008) during the restrictions. This study found that cases of paediatric AGE decreased significantly the during COVID-19 restrictions, suggesting that studies should be conducted to determine whether this reduction was a result of good hand hygiene and social distancing or just a result of altered health-seeking behaviour among children.

Evaluation of Xpert MTB/RIF Assay on Stool Samples for the Diagnosis of Pulmonary Tuberculosis among the Pediatric Population.

Objective Microbiological confirmation of tuberculosis (TB) in pediatric cases is challenging due to its paucibacillary nature and difficulty in specimen collection. This study aimed to validate stool as an alternative sample for the diagnosis of pediatric pulmonary TB via Xpert MTB/RIF (Xpert) assay. Materials and Methods This cross-sectional study included 75 pediatric patients up to 10 years of age with signs and symptoms suggestive of TB. From each recruited patient, pulmonary and stool samples were collected in a sterile container. The collected samples were subjected to Ziehl-Neelsen staining, BACTEC MGIT 960 culture (MGIT), Xpert, and in-house multiplex polymerase chain reaction for TB diagnosis. Results About 13.33% (10/75) of the pulmonary samples and, of them, 50% (5/75) of the stool samples were positive by Xpert assay. The sensitivity and specificity of Xpert assay with stool and pulmonary samples were 50 (95% confidence interval [CI]: 18.71-81.29%) and 100% (95% CI: 94.48-100%), respectively. Conclusion The Xpert assay on stool samples showed limited sensitivity and good specificity in the diagnosis of pulmonary TB. Therefore, it can be proposed as an alternative screening sample to diagnose TB in pediatric cases for which getting a respiratory sample is extremely difficult. However, further studies with greater number of samples and multiple baseline variables are required to support our findings. Strategies to optimize stool Xpert assay should be performed to enhance the sensitivity of this method to detect Mycobacterium tuberculosis in children.

Limitations of PCR detection of filarial DNA in human stools from subjects non-infected with soil-transmitted helminths.

The standard techniques for diagnosis of human filariasis are the microscopic examination of blood smears or skin biopsies, which are relatively invasive and poorly sensitive at low levels of infection. Recently, filarial DNA has been detected in fecal samples from non-human primates in Central Africa. The aim of this study was to demonstrate proof-of-concept of a non-invasive molecular diagnosis technique for human filariasis by targeting fragments of 12S rDNA, Cox1, ITS1 and LL20-15kDa ladder antigen-gene by conventional PCR in DNA extracted from stool samples of 52 people infected with Mansonella perstans and/or Loa loa. Of these, 10 patients were infected with soil-transmitted helminths (Trichuris trichiura and/or Ascaris lumbricoides), and none were positive for Necator americanus. Interestingly, no filarial gene fragments were detected in the stools of any of the 52 patients. Future studies should evaluate whether a co-infection with soil-transmitted helminths causing gastrointestinal bleeding and likely allowing (micro)filaria exit into the digestive tract, may facilitate the molecular detection of filarial DNA fragments in stool samples.

TITLE: Limites de la détection par PCR d'ADN de filaires dans les selles humaines de sujets non-infectés par les géohelminthes. ABSTRACT: Les techniques standards de diagnostic des filarioses humaines (examen microscopique de gouttes épaisses ou de biopsies cutanées) sont relativement invasives et peu sensibles à de faibles niveaux d'infection. De l'ADN de filaires a été récemment détecté dans des échantillons de fèces de primates non-humains en Afrique centrale. L'objectif de cette étude était de démontrer la preuve de concept d'un diagnostic moléculaire non invasif des filarioses chez l'homme en ciblant des fragments d'ADNr 12S, Cox1, ITS1 et l'antigène LL20-15kDa par PCR classique. L'ADN a été extrait d'échantillons de selles de 52 personnes infectées par Mansonella perstans et/ou Loa loa. Parmi ces patients, dix étaient infectés par des géohelminthes (Trichuris trichiura et/ou Ascaris lumbricoides) et aucun n'était positif pour Necator americanus. De manière intéressante, aucun fragment de gène de filaires n'a été détecté dans les selles des 52 patients. Des études futures devraient être menées pour évaluer si une coinfection avec des géohelminthes (provoquant des hémorragies gastro-intestinales et permettant probablement l'effraction de (micro)filaires dans le tube digestif) facilite la détection moléculaire de fragments d'ADN de filaires dans les selles.

Molecular Identification and Characterization of a Genotype 3 Hepatitis E Virus (HEV) Strain Detected in a Wolf Faecal Sample, Italy.

Hepatitis E virus (HEV) infection is a major health problem worldwide. In developed countries, zoonotic transmission of HEV genotypes (Gt) 3 and 4 is caused by the ingestion of raw or undercooked meat of infected pigs and wild boars, the main reservoirs of HEV. However, additional animals may harbour HEV or HEV-related strains, including carnivores. In this study, we investigated the molecular epidemiology of orthohepeviruses in wild canids by screening a total of 136 archival faecal samples, collected from wolves (42) and red foxes (94) in Northwestern Italy. Orthohepevirus RNA was identified in a faecal specimen, collected from a wolf carcass in the province of La Spezia (Liguria Region, Italy). The nearly full-length (7212 nucleotides) genome of the strain HEV/81236/Wolf/2019/ITA (GenBank accession no. MZ463196) was determined by combining a sequence-independent single-primer amplification (SISPA) approach with the Oxford Nanopore Technologies sequencing platform. Upon phylogenetic analysis, the HEV detected in wolf was segregated into clade HEV-3.1, displaying the highest nucleotide (nt) identity (89.0-93.3%) to Gt3 strains belonging to subtype c. Interestingly, the wolf faecal sample also contained porcine astrovirus sequences, endorsing the hypothesis of a dietary origin of the HEV strain due to preying habits.

Cell culture demonstrates superior sensitivity over one step real time RT PCR and nested VP1 amplification for Enteroviruses.

This study evaluated and compared the sensitivity profile of routine cell culture, nested VP1 amplification and one step real time RT PCR for Enteroviruses. Serially diluted spiked samples of four model viruses (EV71, CVA16, CVB5 and PV1) and 32 true positive samples including Poliovirus (PV1 & PV3), Coxsackie virus (CVB5, CVB3, CVB1 & CVA4, 10, 16), Echovirus (Echo 6, 7, 11, 13, 18, 25 & 30) and Enterovirus 71 (E71), and 32 true negative stool samples were subjected to cell culture, nested RT PCR and one step real time RT PCR. The result of sensitivity test indicated superior sensitivity with one step real time RT PCR (75 %, 24/32) against cell culture (71.9 %, 23/32) and nested RT PCR (65.6 %, 21/32). The most specific test was cell culture (100 %, 32/32), followed by nested RT PCR (96.9 %, 31/32). Positive predictive values were 100 %: 23/23, 95.5 %; 21/22 and 88.9 %; 24/27, for cell culture, nested RT PCR and one step real time RT PCR, respectively, and one step real time RT PCR had the highest negative predictive value (78.4 %, 29/37). Overall result indicate relatively high analytical sensitivity with all the tests, suggesting superior performance by cell culture. Therefore, cell culture is the gold standard. However, considering intensive nature of cell cultures and prolong window for results, it is wise to consider one step real time RT PCR in routine diagnosis for its added advantages. Meanwhile, selecting a combination of tests can maximize detection, depending on the laboratory strength.

Fecal Microbiota Transplantation in Patients With Recurrent Clostridium difficile Infection: A Four-Year Single-Center Retrospective Review.

BACKGROUND: Clostridium difficile infection (CDI) is a common cause of hospital and community-acquired diarrhea with an annual incidence of 453,000 cases in the USA. The white race, female gender, and age over 65 years are known risk factors. Recurrence of CDI is a major problem in patients taking antibiotics for prolonged periods. These patients are observed to have reduced diversity of the intestinal microbiome. Fecal microbiota transplantation (FMT) can restore the healthy flora in the gut, thus breaking the cycle of recurrent infection. Our study aimed to analyze the efficacy of FMT and the recurrence of CDI after FMT. We also aimed to investigate the effects of comorbidities on the outcome of FMT. METHODS: After obtaining approval from the institutional review board, we included 64 patients who had received FMT at our institution from October 2015 to November 2019. All the patients over 16 years of age in both inpatient and outpatient settings were included. Patients under 16 years of age and patients treated without FMT were excluded. Frozen stool from a standardized stool bank (OpenBiome) was used. The thawed specimen was instilled into the terminal ileum or the cecum. Patients were followed up for the next 1 year for analysis of improvement in symptoms, recurrence, and repeat FMT. RESULTS: On the 2-months follow-up, 75% of patients reported symptomatic improvement, 15.6% reported no improvement while 9.4% did not follow up. Twenty-six (40.6%) patients had CDI recurrence in the following year; and 69.2% of patients with recurrence underwent a repeat FMT. There was no statistically significant correlation between CDI recurrence and the age (P value = 0.68), gender (P value = 0.61), previous use of proton pump inhibitors (PPIs, P value = 0.11) or antibiotics (P value = 0.45). There was a statistically significant correlation noted with the use of immunosuppressants and recurrence (P value = 0.04). CONCLUSIONS: FMT is a successful treatment modality for refractory and recurrent CDI. Repeat treatments can be beneficial if there is a lack of initial response. Being immunosuppressed with medications is associated with the risk of recurrence.

Methods for detection of Helicobacter pylori from stool sample: current options and developments.

Accurate detection of Helicobacter pylori infection and determination of antibiotics have significant meaning in clinical practice. The detection methods can be categorized into two types, invasive and non-invasive, but nowadays we use the urease breath test most frequently which is non-invasive. However, many developing countries cannot meet the requirements for having specialized equipment and they lack trained personnel. Also, for the children, it is difficult to make them cooperate for the test. Methods that detect Helicobacter pylori from stool sample can be a promising alternative for detection used in children and mass screening. Stool antigen tests have several advantages such as rapidity, simplicity, and cheapness, though their results may be influenced by the heterogenicity of antigens, the nature of biochemical techniques, and the amount of antigen presented in the stool. PCR-based methods can specifically detect Helicobacter pylori infection and antibiotic resistance by targeting specific gene sequence, but they also are limited by the requirements of facilities and experts, the existence of inhibitory substance, and interference from the dead bacteria. Some novel methods also deserve our attention. Here we summarized the results of researches about methods using stool sample and we hope our work can help clinicians choose the appropriate test in clinical practice.