Rapid and multi-target genotyping of Helicobacter pylori with digital microfluidics.

Helicobacter pylori (H. pylori) infection correlates closely with gastric diseases such as gastritis, ulcers, and cancer, influencing more than half of the world's population. Establishing a rapid, precise, and automated platform for H. pylori diagnosis is an urgent clinical need and would significantly benefit therapeutic intervention. Recombinase polymerase amplification (RPA)-CRISPR recently emerged as a promising molecular diagnostic assay due to its rapid detection capability, high specificity, and mild reaction conditions. In this work, we adapted the RPA-CRISPR assay on a digital microfluidics (DMF) system for automated H. pylori detection and genotyping. The system can achieve multi-target parallel detection of H. pylori nucleotide conservative genes (ureB) and virulence genes (cagA and vacA) across different samples within 30 min, exhibiting a detection limit of 10 copies/rxn and no false positives. We further conducted tests on 80 clinical saliva samples and compared the results with those derived from real-time quantitative polymerase chain reaction, demonstrating 100% diagnostic sensitivity and specificity for the RPA-CRISPR/DMF method. By automating the assay process on a single chip, the DMF system can significantly reduce the usage of reagents and samples, minimize the cross-contamination effect, and shorten the reaction time, with the additional benefit of losing the chance of experiment failure/inconsistency due to manual operations. The DMF system together with the RPA-CRISPR assay can be used for early detection and genotyping of H. pylori with high sensitivity and specificity, and has the potential to become a universal molecular diagnostic platform.

Tryptophan Ameliorates Barrier Integrity and Alleviates the Inflammatory Response to Enterotoxigenic Escherichia coli K88 Through the CaSR/Rac1/PLC-γ1 Signaling Pathway in Porcine Intestinal Epithelial Cells.

Background: Impaired intestinal barrier integrity plays a crucial role in the development of many diseases such as obesity, inflammatory bowel disease, and type 2 diabetes. Thus, protecting the intestinal barrier from pathological disruption is of great significance. Tryptophan can increase gut barrier integrity, enhance intestinal absorption, and decrease intestinal inflammation. However, the mechanism of tryptophan in decreasing intestinal barrier damage and inflammatory response remains largely unknown. The objective of this study was to test the hypothesis that tryptophan can enhance intestinal epithelial barrier integrity and decrease inflammatory response mediated by the calcium-sensing receptor (CaSR)/Ras-related C3 botulinum toxin substrate 1 (Rac1)/phospholipase Cγ1 (PLC-γ1) signaling pathway. Methods: IPEC-J2 cells were treated with or without enterotoxigenic Escherichia coli (ETEC) K88 in the absence or presence of tryptophan, CaSR inhibitor (NPS-2143), wild-type CaSR overexpression (pcDNA3.1-CaSR-WT), Rac1-siRNA, and PLC-γ1-siRNA. Results: The results showed that ETEC K88 decreased the protein concentration of occludin, zonula occludens-1 (ZO-1), claudin-1, CaSR, total Rac1, Rho family member 1 of porcine GTP-binding protein (GTP-rac1), phosphorylated phospholipase Cγ1 (p-PLC-γ1), and inositol triphosphate (IP3); suppressed the transepithelial electrical resistance (TEER); and enhanced the permeability of FITC-dextran compared with the control group. Compared with the control group, 0.7 mM tryptophan increased the protein concentration of CaSR, total Rac1, GTP-rac1, p-PLC-γ1, ZO-1, claudin-1, occludin, and IP3; elevated the TEER; and decreased the permeability of FITC-dextran and contents of interleukin-8 (IL-8) and TNF-α. However, 0.7 mM tryptophan+ETEC K88 reversed the effects induced by 0.7 mM tryptophan alone. Rac1-siRNA+tryptophan+ETEC K88 or PLC-γ1-siRNA+tryptophan+ETEC K88 reduced the TEER, increased the permeability of FITC-dextran, and improved the contents of IL-8 and TNF-α compared with tryptophan+ETEC K88. NPS2143+tryptophan+ETEC K88 decreased the TEER and the protein concentration of CaSR, total Rac1, GTP-rac1, p-PLC-γ1, ZO-1, claudin-1, occludin, and IP3; increased the permeability of FITC-dextran; and improved the contents of IL-8 and TNF-α compared with tryptophan+ETEC K88. pcDNA3.1-CaSR-WT+Rac1-siRNA+ETEC K88 and pcDNA3.1-CaSR-WT+PLC-γ1-siRNA+ETEC K88 decreased the TEER and enhanced the permeability in porcine intestine epithelial cells compared with pcDNA3.1-CaSR-WT+ETEC K88. Conclusion: Tryptophan can improve intestinal epithelial barrier integrity and decrease inflammatory response through the CaSR/Rac1/PLC-γ1 signaling pathway.

The Lst Sialyltransferase of Neisseria gonorrhoeae Can Transfer Keto-Deoxyoctanoate as the Terminal Sugar of Lipooligosaccharide: a Glyco-Achilles Heel That Provides a New Strategy for Vaccines to Prevent Gonorrhea.

The lipooligosaccharide (LOS) of Neisseria gonorrhoeae plays key roles in pathogenesis and is composed of multiple possible glycoforms. These glycoforms are generated by the process of phase variation and by differences in the glycosyltransferase gene content of particular strains. LOS glycoforms of N. gonorrhoeae can be terminated with an N-acetylneuraminic acid (Neu5Ac), which imparts resistance to the bactericidal activity of serum. However, N. gonorrhoeae cannot synthesize the CMP-Neu5Ac required for LOS biosynthesis and must acquire it from the host. In contrast, Neisseria meningitidis can synthesize endogenous CMP-Neu5Ac, the donor molecule for Neu5Ac, which is a component of some meningococcal capsule structures. Both species have an almost identical LOS sialyltransferase, Lst, that transfers Neu5Ac from CMP-Neu5Ac to the terminus of LOS. Lst is homologous to the LsgB sialyltransferase of nontypeable Haemophilus influenzae (NTHi). Studies in NTHi have demonstrated that LsgB can transfer keto-deoxyoctanoate (KDO) from CMP-KDO to the terminus of LOS in place of Neu5Ac. Here, we show that Lst can also transfer KDO to LOS in place of Neu5Ac in both N. gonorrhoeae and N. meningitidis Consistent with access to the pool of CMP-KDO in the cytoplasm, we present data indicating that Lst is localized in the cytoplasm. Lst has previously been reported to be localized on the outer membrane. We also demonstrate that KDO is expressed as a terminal LOS structure in vivo in samples from infected women and further show that the anti-KDO monoclonal antibody 6E4 can mediate opsonophagocytic killing of N. gonorrhoeae Taken together, these studies indicate that KDO expressed on gonococcal LOS represents a new antigen for the development of vaccines against gonorrhea.IMPORTANCE The emergence of multidrug-resistant N. gonorrhoeae strains that are resistant to available antimicrobials is a current health emergency, and no vaccine is available to prevent gonococcal infection. Lipooligosaccharide (LOS) is one of the major virulence factors of N. gonorrhoeae The sialic acid N-acetylneuraminic acid (Neu5Ac) is present as the terminal glycan on LOS in N. gonorrhoeae In this study, we made an unexpected discovery that KDO can be incorporated as the terminal glycan on LOS of N. gonorrhoeae by the alpha-2,3-sialyltransferase Lst. We showed that N. gonorrhoeae express KDO on LOS in vivo and that the KDO-specific monoclonal antibody 6E4 can direct opsonophagocytic killing of N. gonorrhoeae These data support further development of KDO-LOS structures as vaccine antigens for the prevention of infection by N. gonorrhoeae.

Identification of bacteria-derived HLA-bound peptides in melanoma.

A variety of species of bacteria are known to colonize human tumours1-11, proliferate within them and modulate immune function, which ultimately affects the survival of patients with cancer and their responses to treatment12-14. However, it is not known whether antigens derived from intracellular bacteria are presented by the human leukocyte antigen class I and II (HLA-I and HLA-II, respectively) molecules of tumour cells, or whether such antigens elicit a tumour-infiltrating T cell immune response. Here we used 16S rRNA gene sequencing and HLA peptidomics to identify a peptide repertoire derived from intracellular bacteria that was presented on HLA-I and HLA-II molecules in melanoma tumours. Our analysis of 17 melanoma metastases (derived from 9 patients) revealed 248 and 35 unique HLA-I and HLA-II peptides, respectively, that were derived from 41 species of bacteria. We identified recurrent bacterial peptides in tumours from different patients, as well as in different tumours from the same patient. Our study reveals that peptides derived from intracellular bacteria can be presented by tumour cells and elicit immune reactivity, and thus provides insight into a mechanism by which bacteria influence activation of the immune system and responses to therapy.

Helicobacter pylori in Pediatric Patients.

Helicobacter pylori causes one of the most common chronic bacterial infections. Clinical manifestations include asymptomatic chronic gastritis, gastric and duodenal ulcers, adenocarcinoma, and gastric mucosa-associated lymphoid tissue lymphoma in adults. In children, most H pylori infections are asymptomatic despite being associated with microscopic gastric inflammation, and children rarely develop complications associated with infection. Due to rising resistance and lack of symptomatic improvement in the absence of peptic ulcer disease, testing and eradication therapy are recommended only for the subset of patients in whom there is a high suspicion of peptic ulcer disease. Studies do not support the role of H pylori infection in functional disorders such as recurrent abdominal pain. A variety of diagnostic modalities exist; therefore, it is important to understand the appropriate approach to diagnosing H pylori infection. The joint European Society for Pediatric Gastroenterology, Hepatology, and Nutrition/North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition guidelines were updated in 2016. Antibiotic and proton pump inhibitor weight-based dosing guidelines have changed to prevent ineffective treatment from increasing antimicrobial resistance. Treatment can also be guided by antibiotic sensitivities obtained from H pylori culture. Patients should be tested again after treatment to confirm eradication.

Review: Diagnosis of Helicobacter pylori infection.

New imaging techniques are still the topic of many evaluations for both the diagnosis of Helicobacter pylori gastritis and the detection of early gastric cancer. Concerning invasive tests, there were studies on the reuse of the rapid urease test material for other tests, and a novel fluorescent method to be used for histology but with limited sensitivity. Progress occurred essentially in the molecular methods area, especially next-generation sequencing which is applied to detect both H pylori and the mutations associated with antibiotic resistance. For non-invasive tests, a few studies have been published on the validity of breath collection bags, the shortening of the testing time, the performance of different analysers or the added value of citric acid in the protocol. The accuracy of serological immunochromatographic tests is also improving. Multiplex serology detecting antibodies to certain proteins allows confirmation of a current infection. Dried blood spots can be used to collect and store blood without a loss of accuracy. Finally, the serum antibody titer can be useful in predicting the risk of gastric cancer. Several stool antigen tests were evaluated with good results, and a novel test using immunomagnetic beads coated with monoclonal antibodies is potentially interesting. PCR detection in stools can also be effective but needs an efficient DNA extraction method. The use of easyMAG® (bioMérieux) combined with Amplidiag® H pylori + ClariR (Mobidiag) appears to be powerful.

Diagnosis of Helicobacter pylori infection in the elderly using an immunochromatographic assay-based stool antigen test.

The diagnostic value of Helicobacter pylori stool antigen (HpSA) tests in elderly subjects remains unclear. The objective of this study was to assess the diagnostic accuracy of the immunochromatographic assay-based HpSA test in a male elderly cohort and identify factors affecting the accuracy. Data for asymptomatic elderly male citizens (≥65 years old) who received health checkups at the Chinese PLA General Hospital between July 2007 and November 2018 were collected. The diagnostic accuracy of the HpSA test was determined using the 13 C-urea breath test as a reference standard. Associations between baseline comorbidities and the accuracy of the HpSA test were analyzed. In total, 316 participants were enrolled, including 193 in the pre-treatment group (77.2 ± 7.8 years old) and 123 in the post-treatment group (78.7 ± 8.3 years old). The accuracy (91.5%, 91.2%, and 91.9%) and specificity (97.6%, 98.7%, and 96.0%) were high in all participants, pre- and post-treatment groups, respectively. However, sensitivities were only 68.7%, 65.1%, and 75.0%, respectively. In the pre-treatment group, constipation was associated with decreased sensitivity (p = 0.039), while colorectal polyps were associated with increased sensitivity (p = 0.010). Multivariate analysis indicated that constipation and colorectal polyps are independent factors for the sensitivity of HpSA in the pre-treatment group. The immunochromatographic assay-based HpSA test achieved high accuracy with high specificity but suboptimal sensitivity in the elderly male cohort. Constipation and colorectal polyps were negatively and positively associated with HpSA sensitivity, respectively, in the pre-treatment group.

Helicobacter pylori Antigen But Not the Organism Is Occasionally Present Within Germinal Centers: Implications for Patient Management and Biology.

Helicobacter pylori infection is present in two thirds of the world's population and induces a myriad of human diseases, ranging from gastritis to gastric adenocarcinoma and mucosa-associated lymphoid tissue lymphoma. Detection is critical for treatment and may require immunohistochemical (IHC) staining when organisms are not visible on hematoxylin and eosin. We have encountered cases in which IHC for Helicobacter pylori failed to demonstrate curvilinear or coccoid organisms, but did show a reticular pattern of immunoreactivity involving the underlying germinal centers. We performed a systematic retrospective evaluation of the frequency of H. pylori germinal center immunoreactivity over a 54-month period through evaluation of 367 gastric specimens. H. pylori germinal center immunoreactivity was observed in 5% of cases with germinal centers. Nine of 11 (81%) patients with H. pylori germinal center immunoreactivity had concurrent or recent H. pylori infection, in comparison to 36% of patients with germinal centers present but no immunoreactivity (n=9 of 25 patients, P=0.03). None of the patients with germinal center immunoreactivity developed mucosa-associated lymphoid tissue lymphoma. In situ hybridization for H. pylori performed on 3 cases with positive germinal center IHC was negative for H. pylori nucleic acids within those germinal centers, demonstrating that only the antigen is present. This work demonstrates that H. pylori antigen, but not viable organisms, is present in germinal centers in 5% of gastric specimens, and is associated with recent or concurrent H. pylori infection. We advocate for reporting of all H. pylori germinal center immunoreactivity with a recommendation for ancillary H. pylori testing.

A combination of iron metabolism indexes and tuberculosis-specific antigen/phytohemagglutinin ratio for distinguishing active tuberculosis from latent tuberculosis infection.

BACKGROUND: Discriminating active tuberculosis (ATB) from latent tuberculosis infection (LTBI) remains challenging. This study aimed to investigate a diagnostic model based on a combination of iron metabolism and the TB-specific antigen/phytohemagglutinin ratio (TBAg/PHA ratio) in T-SPOT.TB assay for differentiation between ATB and LTBI. METHODS: A total of 345 participants with ATB (n=191) and LTBI (n=154) were recruited based on positive T-SPOT.TB results at Tongji hospital between January 2017 and January 2020. Iron metabolism analysis was performed simultaneously. A diagnostic model for distinguishing ATB from LTBI was established according to multivariate logistic regression. RESULTS: The TBAg/PHA ratio showed 64.00% sensitivity and 90.10% specificity in distinguishing ATB from LTBI when a threshold of 0.22 was used. All iron metabolism biomarkers in the ATB group were significantly different from those in the LTBI group. Specifically, serum ferritin and soluble transferrin receptor in ATB were significantly higher than LTBI. On the contrary, serum iron, transferrin, total iron binding capacity, and unsaturated iron binding capacity in ATB were significantly lower than LTBI. The combination of iron metabolism indicators accurately predicted 60.00% of ATB cases and 91.09% of LTBI subjects, respectively. Moreover, the combination of iron metabolism indexes and TBAg/PHA ratio resulted in a sensitivity of 88.80% and specificity of 90.10%. Furthermore, the performance of models established in the Qiaokou cohort was confirmed in the Caidian cohort. CONCLUSIONS: The data suggest that the combination of iron metabolism indexes and TBAg/PHA ratio could serve as a biomarker to distinguish ATB from LTBI in T-SPOT-positive individuals.

Clinical Performance of the Automated LIAISON® Meridian H. pylori SA Stool Antigen Test.

BACKGROUND: Antigens derived from Helicobacter pylori can be used as stool biomarkers to assist in the diagnosis of H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian. METHODS: This prospective multisite study enrolled patients undergoing an esophagogastroduodenoscopy with collection of biopsy and stool specimens. Adult patients (≥22 years) participated in the study from February 2017 to August 2018. Specimens of the stomach were tested by three methods, known as the Composite Reference Method: (1) histological evaluation, (2) culture of the organism, and (3) rapid urease detection test. H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian. RESULTS: 277 patients (63% female) were included in the study. The prevalence of infected subjects was 24.2% in this study cohort. Clinical performance assessed against the Composite Reference Method showed very good agreement (Cohen's kappa = 0.922), with good sensitivity (95.5%) and specificity (97.6%). Reproducibility study results showed total imprecision ranging from 3.1% to 13.9% CV. CONCLUSION: The automated LIAISON® Meridian H. pylori SA assay brings reliable noninvasive testing for H. pylori to the laboratory that is in very good agreement with the current, more invasive biopsy-based methods such as histology, culture, or rapid urease test. The clinical trial identifiers are NCT03060746 (pretherapy) and NCT03060733 (posttherapy).H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian H. pylori infection. Since current assays have variable performance, we assessed the clinical performance of the automated LIAISON® Meridian.

Helicobacter pylori: A Review of Current Diagnostic and Management Strategies.

As one of the most prevalent infections globally, Helicobacter pylori (H. pylori) continues to present diagnostic and therapeutic challenges for clinicians worldwide. Diagnostically, the "test-and-treat" strategy is the recommended approach for healthcare practitioners when managing this potentially curable disease. The choice of testing method should be based on several factors including patient age, presenting symptoms, and medication use, as well as test reliability, availability, and cost. With rising antibiotic resistance, particularly of macrolides, care must be taken to ensure that therapy is selected based on regional resistance patterns and prior antibiotic exposure. In the USA, macrolide antibiotic resistance rates in some areas have reached or exceeded a generally accepted threshold, such that clarithromycin triple therapy may no longer be an appropriate first-line empiric treatment. Instead, bismuth quadruple therapy should be considered, while levofloxacin-based or alternative macrolide-containing therapies are also options. Once treated, it is essential to test for eradication as untreated H. pylori is associated with serious complications including peptic ulcer disease, mucosa-associated lymphoid tissue lymphoma, and gastric cancer. This review article aims to consolidate current knowledge of H. pylori infection with a particular emphasis on diagnostic and treatment strategies.

Is the Utilization of Helicobacter pylori Stool Antigen Tests Appropriate in an Urban Canadian Population?

OBJECTIVES: Helicobacter pylori stool antigen test (HpSAT) appropriateness was investigated by assessing its testing and positivity rates in Calgary, Canada. METHODS: The laboratory information system was accessed for all patients who received an HpSAT in 2018. Testing volume, test results, age, and sex of patients were collected. Sociodemographic risk factors and geospatial analysis were performed by matching laboratory data to the 2016 census data. Testing appropriateness was defined as a concordance between testing and positivity rates for each sociodemographic variable. RESULTS: In 2018, 25,518 H pylori stool antigen tests were performed in Calgary, with an overall positivity rate of 14.7%. Geospatial mapping demonstrated significant distribution variations of testing and positivity rates of HpSAT in the city. Certain sociodemographic groups studied (eg, recent immigrants) appeared to be appropriately tested (testing rate relative risk [RR] = 2.26, positivity rate RR = 4.32; P < .0001), while other groups (eg, male) may have been undertested (testing rate RR = 0.85, positivity rate RR = 1.14; P < .0001). CONCLUSIONS: Determining concordance of testing and positivity rate of a laboratory test can be used for assessing testing appropriateness for other diseases in other jurisdictions. This study demonstrated some at-risk patients may be missed for H pylori testing.

Prospective, study comparing the accuracy of two different stool antigen tests (Premier Platinum HpSA and novel ImmunoCard STAT! rapid test) for the diagnosis of Helicobacter pylori infection.

BACKGROUND: At present only monoclonal EIA (enzyme-immunoassay) stool antigen-tests have obtained optimal accuracy in the diagnosis of Helicobacter pylori. Our aim was to evaluate the accuracy of two stool antigen-tests, the validated Premier Platinum HpSA PLUS (EIA test) and the newly available ImmunoCard STAT! HpSA HD (rapid test) for the initial diagnosis and the confirmation of eradication of H. pylori infection. PATIENTS AND METHODS: Patients with indication of H. pylori diagnosis, or confirmation after treatment were included. Data were coded to protect personal data and ensure blindness between tests. Accuracy was considered as coincident diagnosis with the gold standard (13C-urea breath test, UBT). The EIA was used as a bench standard. All stool tests were performed in duplicate. RESULTS: 264 patients completed the protocol (100 naïve, 164 post-eradication). Average age was 52 years, 61% women, 11% ulcer. Positive diagnoses by UBT were 41% for naïve and 17% for post-eradication. Overall ImmunoCard and EIA accuracies were respectively 91% (95%C.I.=88-94%) and 89% (86-93%), sensitivities 72% (67-78%) and 72% (67-78%), and specificities 98% (96-100%), and 95% (92-97%). Concordance between ImmunoCard and EIA was 95% (93-98%). DISCUSSION: Our results indicate that the newly available ImmunoCard rapid stool antigen-test achieves 90% accuracy, with high specificity but suboptimal sensitivity. The ImmunoCard attained equivalent accuracies as the EIA bench standard, with 95% concordance.

Helicobacter pylori infection and gastroduodenal lesions in patients with systemic lupus erythematosus.

OBJECTIVE: The aim of this study was to determine the frequency of Helicobacter pylori in SLE patients and to compare clinical characteristics and gastroduodenal lesions in patients with and without H. pylori infection. METHODS: Adult SLE patients were selected and subjected to endoscopy. Gastroduodenal lesions were examined by endoscopy and biopsy (antrum and corpus). Biopsies were evaluated by hematoxylin and eosin and Giemsa staining. Immunochromatographic membrane-based assay using amplification was used to test for H. pylori antigen (coproantigen) in stool samples in all participants. Clinical characteristics and gastroduodenal lesions were compared between patients with and without H. pylori infection. RESULTS: A total of 118 SLE patients were included (mean age 44.7 ± 11.7 years, mean disease duration 11.6 ± 6.0 years), of whom 101 (85.6%) were receiving non-steroidal anti-inflammatory drugs (NSAIDs). The coproantigen test was positive in 32 (27.1%) patients. H. pylori was present in twenty six patients (22.0%) in the gastric biopsy. The frequency of gastric erosions and gastric ulcers were 55.1% and 0.8%, respectively. Gastric erosions were less frequent in SLE patients with H. pylori infection than those without H. pylori (43.5.7% vs. 62.5%; p = 0.04). The age, disease duration, disease activity, chronic damage, gastroprotective drugs, and immunosuppressive therapy did not differ between the two groups. CONCLUSIONS: We found a high frequency of H. pylori infection in SLE patients. The severity of SLE and reception of gastroprotective therapy do not seem to be related to H. pylori infection. Immunosuppressive therapy may not be protective against H. pylori infection in SLE patients.Key Points• In patients with systemic lupus erythematosus (SLE), the frequency of Helicobacter pylori infection was 39% and gastric erosions were frequent.• Disease activity, chronic damage, gastroprotective drugs, and immunosuppressive therapy may not affect the prevalence of H. pylori infection in SLE patients.

La derivación protocolizada a endoscopía asociada a la detección de Helicobacter pylori mediante antígeno en deposiciones disminuye lista de espera para endoscopía y optimiza la detección de lesiones pre-malignas y cáncer gástrico incipiente / Protocolized referral to endoscopy and Helicobacter pylori detected in stools aimed to decrease endoscopy waiting lists

Background Chile has one of the highest mortality rates by gastric cancer (GC) worldwide. Primary prevention of GC and detection of pre-neoplastic and early neoplastic lesions should be a national priority. Aim To assess the impact of the protocolization of endoscopy referral and the use of H. pylori stool antigen test (HPSA) in the management of dyspepsia to decrease the waiting list for endoscopy and increase the detection of gastric pre-neoplastic and early neoplastic lesions. Material and Methods We included all patients referred to the Endoscopy Unit of a regional hospital, from January 2015 to December 2017. We also included patients with known pre-neoplastic lesions and all those with first degree relatives with GC. We implemented protocols for referral of patients with dyspepsia considering the use of HPSA test, prioritizing to endoscopy those with a higher risk of GC. Results A total of 4,641 endoscopies and 2,631 HPSA tests were carried out. After the adoption of these protocols, we observed a 52% decrease in the waiting time for endoscopy. The GC detection rate in this period was 1.8 to 3.1 cases per 100 endoscopies. After the adoption of the protocols, we observed a significant increase in early GC detection rate (from none in 2015 to 13% in 2017, p = 0.03). Conclusions The protocolization of the referral for endoscopy associated with widespread use of HPSA test in the management of patients with dyspepsia, are successful strategies to decrease waiting lists for endoscopy and optimize the detection rate of pre-neoplastic lesions and early GC.

A case with neurological abnormalities caused by Rickettsia raoultii in northwestern China.

BACKGROUND: The number of new rickettsial species are rapidly increasing, and increasing numbers of Rickettsia raoultii (R. raoultii) infection cases have been detected in humans. However, neurological abnormalities caused by R. raoultii are rarely reported, especially in northwestern China. CASE PRESENTATION: A 36-year-old Kazakh shepherd with an attached tick on part temporalis, presented with right eyelid droop, lethargy, fever, headache, fever (38.0-41.0 °C) and erythematous rash. The examination of cerebrospinal fluid (CSF) showed cerebrospinal pressure of 200 mm H2O, leukocyte count of 300.0 × 106/L, adenosine deaminase of 2.15 U/L, and total protein concentration of 0.93 g/L. The diagnosis of R. raoultii infection was confirmed by six genetic markers, and semi-quantified by enzyme-linked immunosorbent assay for rickettsial antigen. The patient gradually recovered after treatment with doxycycline and ceftriaxone. R. raoultii DNA was found both in a tick detached from this patient and in 0.18% (2/1107) of blood samples collected from local shepherds. CONCLUSIONS: This is the first reported case with neurological abnormalities caused by R. raoultii in northwestern China. It is vital to detect rickettsial agents both in blood and CSF for tick bite patients with neurological abnormalities. Public health workers and physicians should pay attention to neurological abnormalities caused by Rickettsia.

Hydrogen Peroxide Production of Group A Streptococci (GAS) is emm-Type Dependent and Increased at Low Temperatures.

Group A streptococcus (GAS) is an important human pathogen whose clinical isolates differ in their ability to produce hydrogen peroxide (H2O2). H2O2 is primarily produced by the enzyme lactate oxidase (LctO), an in depth in silico research revealed that all genome-sequenced GAS possess the required gene lctO. The importance of lctO for GAS is underlined by its highly conserved catabolite control element (cre box) as well as its perfect promotor sequence in comparison to the known consensus sequences of the Gram-positive model organism Bacillus subtilis. In this study, we provide further insight in the function and regulation of lactate oxidase by analyzing a large group of clinical GAS isolates. We found that H2O2 production increased over time in the late stationary phase; after 4 days of incubation, 5.4% of the isolates showed a positive result at 37 °C, while the rate increased to 16.4% at 20 °C. This correlation between H2O2 production and low temperatures suggests additional regulatory mechanisms for lctO besides catabolite control protein A (CcpA) and indicates that lctO might play a role for GAS energy metabolism at sub-body temperatures. Furthermore, we could identify that H2O2 production was different among clinical isolates; we could correlate H2O2 production to emm-types, indicating that emm-types 6 and 75 had the highest rate of H2O2 production. The emm-type- and temperature-dependent H2O2 production of clinical GAS isolates might contribute to their different survival strategies.

A Clinical Diagnostic Dilemma: a Case Report of Primary Hepatic Tuberculosis.

BACKGROUND: Primary hepatic tuberculosis is a very rare clinical form of tuberculosis, with atypical clinical presentations and nonspecific imaging features. This results in great difficulty to reach a correct and timely clinical diagnosis. Traditionally, liver biopsy is the gold standard for its diagnosis. Here we assessed the effectiveness of using a T-SPOT.TB test in the early diagnosis of primary hepatic tuberculosis. METHODS: We report a case of primary hepatic tuberculosis whose location of hepatic lesion renders it hard to perform a biopsy. Instead, a T-SPOT.TB test was utilized to help in the early diagnosis of this rare form of tuberculosis. A conventional fourdrug regimen for anti-tubercular therapy together with vitamin B6 was initiated and maintained for 6 months. RESULTS: The T-SPOT.TB test was highly positive for ESAT-6 (87 > 20) and CFP-10 (89 > 20). Dull pain in the upper right abdomen was gone 2 months post treatment. The abnormal lesions shown in an MRI reduced significantly 4 months post treatment. Spot count for ESAT-6 and CFP-10 decreased 6 months post treatment. CONCLUSIONS: The results of this study suggest the critical role of T-SPOT.TB test in the earlier diagnosis of prima¬ry hepatic tuberculosis for those patients who have difficulties having a hepatic biopsy.

GastroPanel® Biomarker Assay: The Most Comprehensive Test for Helicobacter pylori Infection and Its Clinical Sequelae. A Critical Review.

BACKGROUND/AIM: Several clinical conditions seriously hamper the diagnostic accuracy of the commonly used tests for Helicobacter pylori (Hp), 13C-urea breath test (UBT) and stool antigen test (SAT). The present communication is a critical review of the potential limitations of UBT and SAT, and describes the approach on how these can be avoided. Drawbacks of the Hp tests: False-negative results are most often due to low bacterial load in the stomach due to: i) use of proton pump inhibitor medication; ii) use of antibiotics; iii) presence of atrophic gastritis and hypoacid stomach; iv); bleeding peptic ulcer; v) gastric cancer (GC) and vi) mucosal-associated lymphatic tissue lymphoma. The UBT also gives false-positive results when urease-producing bacterial species, other than Hp colonize an acid-free stomach. Importantly, neither UBT nor SAT are capable of diagnosing atrophic gastritis, thus missing the patients at highest risk for GC. GastroPanel® (Biohit Oyj, Finland) circumvents these shortcomings with a serological test consisting of a panel of stomach-specific biomarkers: pepsinogen I, pepsinogen II, gastrin-17 and Hp antibodies. GastroPanel® is a tool for non-invasive examination of i) dyspeptic patients for exclusion or diagnosis of Hp or atrophic gastritis, also disclosing the status of gastric acid output; ii) for screening of asymptomatic individuals at risk of GC; and iii) for comprehensive diagnosis of Hp infection. GastroSoft® application integrates the biomarker profile with the patient's medical information, accurately classifying the biomarker profiles into eight diagnostic categories. CONCLUSION: Given that Hp is the single most important risk factor of GC, the non-invasive diagnosis and screening of Hp should be based on more accurate and more comprehensive testing than UBT or SAT alone. The GastroPanel® is such test, being completely devoid of the known serious shortcomings of UBT and SAT.