Delfina C. Dominguez

Calcium signalling in bacteria

Whereas the importance of calcium as a cell regulator is well established in eukaryotes, the role of calcium in prokaryotes is still elusive. Over the past few years, there has been an increased interest in the role of calcium in bacteria. It has been demonstrated that as in eukaryotic organisms, the intracellular calcium concentration in prokaryotes is tightly regulated ranging from 100 to 300 nM. It has been found that calcium ions are involved in the maintenance of cell structure, motility, transport and cell differentiation processes such as sporulation, heterocyst formation and fruiting body development. In addition, a number of calcium‐binding proteins have been isolated in several prokaryotic organisms. The characterization of these proteins and the identification of other factors suggest the possibility that calcium signal transduction exists in bacteria. This review presents recent developments of calcium in bacteria as it relates to signal transduction.

A Versatile PDMS/Paper Hybrid Microfluidic Platform for Sensitive Infectious Disease Diagnosis

Bacterial meningitis is a serious health concern worldwide. Given that meningitis can be fatal and many meningitis cases occurred in high-poverty areas, a simple, low-cost, highly sensitive method is in great need for immediate and early diagnosis of meningitis. Herein, we report a versatile and cost-effective polydimethylsiloxane (PDMS)/paper hybrid microfluidic device integrated with loop-mediated isothermal amplification (LAMP) for the rapid, sensitive, and instrument-free detection of the main meningitis-causing bacteria, Neisseria meningitidis (N. meningitidis). The introduction of paper into the microfluidic device for LAMP reactions enables stable test results over a much longer period of time than a paper-free microfluidic system. This hybrid system also offers versatile functions, by providing not only on-site qualitative diagnostic analysis (i.e., a yes or no answer), but also confirmatory testing and quantitative analysis in laboratory settings. The limit of detection of N. meningitidis is about 3 copies per LAMP zone within 45 min, close to single-bacterium detection sensitivity. In addition, we have achieved simple pathogenic microorganism detection without a laborious sample preparation process and without the use of centrifuges. This low-cost hybrid microfluidic system provides a simple and highly sensitive approach for fast instrument-free diagnosis of N. meningitidis in resource-limited settings. This versatile PDMS/paper microfluidic platform has great potential for the point of care (POC) diagnosis of a wide range of infectious diseases, especially for developing nations.

Calcium binding proteins and calcium signaling in prokaryotes

With the continued increase of genomic information and computational analyses during the recent years, the number of newly discovered calcium binding proteins (CaBPs) in prokaryotic organisms has increased dramatically. These proteins contain sequences that closely resemble a variety of eukaryotic calcium (Ca(2+)) binding motifs including the canonical and pseudo EF-hand motifs, Ca(2+)-binding β-roll, Greek key motif and a novel putative Ca(2+)-binding domain, called the Big domain. Prokaryotic CaBPs have been implicated in diverse cellular activities such as division, development, motility, homeostasis, stress response, secretion, transport, signaling and host-pathogen interactions. However, the majority of these proteins are hypothetical, and only few of them have been studied functionally. The finding of many diverse CaBPs in prokaryotic genomes opens an exciting area of research to explore and define the role of Ca(2+) in organisms other than eukaryotes. This review presents the most recent developments in the field of CaBPs and novel advancements in the role of Ca(2+) in prokaryotes.

Greater Than 95% Success with 14-day Bismuth Quadruple Anti- Helicobacter pylori Therapy: A Pilot Study in US Hispanics

A combination capsule of bismuth, metronidazole, and tetracycline plus omeprazole given as 10‐day therapy has an overall effectiveness of 92–93% in per‐protocol analysis (Grade B) with eradication of 86–91% of metronidazole‐resistant Helicobacter pylori. This study aimed to explore whether extending the duration to 14 days would improve overall effectiveness per protocol to ≥95% (Grade A) in a population in which metronidazole resistance was anticipated to exist.

Evaluation of a novel stool native catalase antigen test for Helicobacter pylori infection in asymptomatic North American children.

Rapid immunochromatographic tests for Helicobacter pylori infection have been developed to allow “near-patient” testing. We therefore performed a pilot study to test a rapid immunochromatographic stool antigen test for the diagnosis of H pylori infection in asymptomatic children. We tested stool specimens collected from children participating in a cohort study in the United States and Mexico. H pylori–positive status was defined by positivity on at least 2 tests: a commercial H pylori stool antigen enzyme immunoassay, an immunoglobulin G antibody enzyme immunoassay, and the 13C-urea breath test. Negative H pylori status was defined by negative findings of all of these tests. Of 52 children (22 girls, 30 boys) 25 were H pylori–positive, 19 H pylori–negative, and 8 uncertain (eg, presumably negative; positive findings on 1 of the 3 noninvasive tests). The sensitivity and specificity of the new stool antigen test for those with definite H pylori status were 100% (exact 95% CI 86.3%–100% and 82.4%–100%, respectively). This rapid stool antigen test may prove useful for point-of-care testing and epidemiological field studies. Larger prospective studies are needed in symptomatic and asymptomatic children for more precise estimates.

Multiplexed instrument-free meningitis diagnosis on a polymer/paper hybrid microfluidic biochip

Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae), and Haemophilus influenzae type b (Hib) are three most common pathogens accounting for most bacterial meningitis, a serious global infectious disease with high fatality, especially in developing nations. Because the treatment and antibiotics differ among each type, the identification of the exact bacteria causing the disease is vital. Herein, we report a polymer/paper hybrid microfluidic biochip integrated with loop-mediated isothermal amplification (LAMP) for multiplexed instrument-free diagnosis of these three major types of bacterial meningitis, with high sensitivity and specificity. Results can be visually observed by the naked eye or imaged by a smartphone camera under a portable UV light source. Without using any specialized laboratory instrument, the limits of detection of a few DNA copies per LAMP zone for N. meningitidis, S. pneumoniae and Hib were achieved within 1h. In addition, these three types of microorganisms spiked in artificial cerebrospinal fluid (ACSF) were directly detected simultaneously, avoiding cumbersome sample preparation procedures in conventional methods. Compared with the paper-free non-hybrid microfluidic biochip over a period of three months, the hybrid microfluidic biochip was found to have a much longer shelf life. Hence, this rapid, instrument-free and highly sensitive microfluidic approach has great potential for point-of-care (POC) diagnosis of multiple infectious diseases simultaneously, especially in resource-limited settings.

Helicobacter pylori Eradication and Change in Markers of Iron Stores Among Non―iron-deficient Children in El Paso, Texas: An Etiologic Intervention Study

Objectives: We assessed whether Helicobacter pylori eradication was followed by changes in iron stores among non–iron-deficient children. Materials and Methods: Double-blind randomized intervention trial on 110 asymptomatic 3- to 10-year-olds with H pylori infection assigned to any of the following 4 arms: both quadruple eradication and iron supplementation, either quadruple sequential eradication or iron supplementation, or placebo only. Hemoglobin, transferrin saturation, and serum ferritin were measured at baseline and 8 months later to assess changes according to study arm, H pylori infection status at ≥45 days, and cytotoxin-associated gene product A status. Results: Intent-to-treat (n = 110) and per-protocol (n = 90) analyses revealed no differences across study arms in changes of iron stores. However, we found that those who had their infection eradicated had a 3-fold increased average change from baseline serum ferritin compared with that of children who remained infected (P < 0.05). Eradication of infection by cytotoxin-associated gene product A negative strains was associated with a larger ferritin increase. Conclusions: In this double-blind randomized trial, the first among non–iron-deficient, asymptomatic H pylori–infected children living in the contiguous United States, we found no effect of H pylori eradication regarding changes in iron stores. However, those who had their infection eradicated at follow-up had a significantly larger increase in serum ferritin from baseline.

Is human herpesvirus 6 linked to Kikuchi-Fujimoto disease? The importance of consistent molecular and serologic analysis

Background Kikuchi-Fujimoto disease, also called histiocytic necrotizing lymphadenitis, is a self-limiting lymphadenopathy of unknown cause. The patient often presents with fever, malaise, and lymphadenopathy primarily involving the cervical lymph nodes. Laboratory findings are nonspecific, and the diagnosis is established by identifying characteristic pathologic features from lymph node biopsy. The etiologic agent has been proposed to be of viral origin; specifically, human herpesvirus 6 (HHV-6) has been implicated. Methods The clinical, histologic, serologic, and molecular data from 471 cases of Kikuchi-Fujimoto disease reported between 1982 and 2001 and their relationship to HHV-6 were reviewed. Results Thirty-two of the 471 patients in this review were tested for HHV-6 serology; all 32 had elevated titers for HHV-6. One hundred twelve patients with lymphadenopathies, including histiocytic necrotizing lymphadenitis, tested positive for HHV-6 by polymerase chain reaction. In situ hybridization results were positive in 41 cases. Conclusion This review illustrates the necessity for consistent molecular and serologic analysis.

Double-blind randomized trial of quadruple sequential Helicobacter pylori eradication therapy in asymptomatic infected children in El Paso, Texas.

Objectives: We assessed the efficacy of a novel quadruple sequential 10-day eradication therapy, its compliance, and reported adverse events in a sample of asymptomatically Helicobacter pylori–infected children in El Paso, Texas, as part of a study aiming to assess the influence of this infection on the levels of markers of iron stores. Patients and Methods: Using a double-blind randomized trial design, 110 asymptomatic children ages 3 to 11 with H pylori infection were randomly assigned to receive either a 10-day course of sequential eradication therapy plus 6 weeks of iron supplementation, eradication therapy plus placebo, iron supplementation plus placebo, or placebo only. H pylori infection status was assessed ≥45 days after treatment using the urea breath test. Analyses compared the proportion of subjects cured according to assignment to and completion of the sequential eradication therapy. Results: Intent-to-treat and per-protocol analyses indicated that 44.3% and 52.9%, respectively, of the children receiving the novel quadruple sequential therapy had their infection eradicated compared with 12.2% and 15.4% in the arms receiving iron or placebo only, respectively (P < 0.001 in both analyses). Study medications were taken with no or only mild adverse events in most children. Conclusions: A quadruple sequential regimen eradicated H pylori in only half the asymptomatic children receiving this treatment. There was no difference in the cure rates of those receiving iron supplementation and those receiving placebo.

Study of methicillin-resistant Staphylococcus aureus on the U.S./Mexico border

BACKGROUND El Paso, Texas and Ciudad Juarez, Mexico comprise the largest U.S./Mexico border population. METHODS Bacterial samples were collected from two hospitals in El Paso and two in Ciudad Juarez and transported to a reference microbiology laboratory in El Paso for microbial identification and antimicrobial susceptibility testing according to NCCLS standards. The presence of the MecA gene, and the prevalence of both the SSCmec IV element and the Panton-Valentine leukocidin were investigated by PCR in all MRSA isolates. RESULTS A total of 201 isolates in El Paso and 128 in Ciudad Juarez of Staphylococcus aureus were identified, of those, MRSA were significantly more prevalent in El Paso than in Ciudad Juarez [89 (44.3%) versus 10 (7.8%) respectively (p<0.0001)]. Thirty one (35%) of MRSA strains isolated in El Paso were community associated. CONCLUSION Significantly higher prevalence of MRSA infections was documented in El Paso compared to Ciudad Juarez.