Laboratory Of Infectious Diseases Research Articles

The virucidal effects of 405\xa0nm visible light on SARS-CoV-2 and influenza A virus

The germicidal potential of specific wavelengths within the electromagnetic spectrum is an area of growing interest. While ultra-violet (UV) based technologies have shown satisfactory virucidal potential, the photo-toxicity in humans coupled with UV associated polymer degradation limit their use in occupied spaces. Alternatively, longer wavelengths with less irradiation energy such as visible light (405 nm) have largely been explored in the context of bactericidal and fungicidal applications. Such studies indicated that 405 nm mediated inactivation is caused by the absorbance of porphyrins within the organism creating reactive oxygen species which result in free radical damage to its DNA and disruption of cellular functions. The virucidal potential of visible-light based technologies has been largely unexplored and speculated to be ineffective given the lack of porphyrins in viruses. The current study demonstrated increased susceptibility of lipid-enveloped respiratory pathogens of importance such as SARS-CoV-2 (causative agent of COVID-19) and influenza A virus to 405 nm, visible light in the absence of exogenous photosensitizers thereby indicating a potential alternative porphyrin-independent mechanism of visible light mediated viral inactivation. These results were obtained using less than expected irradiance levels which are considered safe for humans and commercially achievable. Our results support further exploration of the use of visible light technology for the application of continuous decontamination in occupied areas within hospitals and/or infectious disease laboratories, specifically for the inactivation of respiratory pathogens such as SARS-CoV-2 and Influenza A.

Антенатальная и ранняя постнатальная этиологическая верификация актуальных врожденных вирусных инфекций

Nonspecificity of clinical, laboratory and instrumental manifestations of congenital infectious diseases, including viral infections, and the diversity of methods for etiological verification of pathogens define both the need to choose the optimal approaches to the diagnosis of this pathology, and the feasibility of testing for a broad range of etiologic agents in case of suspected congenital viral infection. The analysis of current guidelines, international consensus documents issued by specialists, and published results of some studies has shown that identification of the genetic material of the pathogen with the use of amniocentesis/ cordocentesis (for cytomegalovirus and parvovirus infections) or in the birth canal (for herpes simplex infection) is the key method for antenatal etiological verification of the widespread viral infections. During the postnatal period, molecular genetic testing is combined with serological diagnosis involving determining specific immunoglobulins M and G, as well as their avidity index.

Counting and identification of molds and yeasts in dry salted shrimp commercialized in Rio Branco, Acre, Brazil

Dry salted shrimp is a popular food in Acre cuisine, especially as a main ingredient in “tacacá” and “rabada no tucupi”. However, different intrinsic and extrinsic factors can result in accelerated deterioration and microbial proliferation, causing food poisoning to the consumer. The objective was to perform the counting and identification of molds and yeasts in dry salted shrimp in establishments that sell typical foods in the municipality of Rio Branco, state of Acre. Shrimp samples from six establishments were collected in sterile bottles, identified, refrigerated and transported to the Laboratory of Infectious Diseases of Animals at the Federal University of Acre, for the manual counting of colony-forming units and identification of fungal genera. As a result, the number of molds and yeasts in the dried salted shrimp samples varied between 1.0x102 and 1.02x106 CFU/g. Nine fungal genera were identified: Aspergillus, Penicillium, Cladosporium, Malassezia, Fusarium, Exophiala, Candida, Curvularia and Trichosporon. The most frequent were Aspergillus, Penicillium and Cladosporium, these being potential producers of mycotoxins. The presence of molds and yeasts in the samples of dried salted shrimp shows the need for greater hygienic-sanitary rigidity of this fish at different stages of manipulation and processing as preventative measures for public health.

Infectious disease screening in a dedicated primary care clinic for children in foster care

BackgroundChildren in foster care are considered at high risk for infectious diseases, and guidelines recommend screening for tuberculosis, hepatitis B and C, syphilis, gonorrhea and chlamydia. Little is known about the prevalence of infectious disease in children in foster care. ObjectivesDescribe infectious disease screening practices in a primary care clinic dedicated to caring for children in foster care. Participants and settingPatients evaluated at a foster care primary care clinic at a southwestern academic center. MethodsRetrospective chart review. ResultsFrom January 1, 2017 through December 31, 2018, 2868 unique patients were evaluated (53 % male, 41 % white, 30 % black, 19 % Hispanic); 1638 (57 %) had any infectious disease laboratory screening done. About 50 % of children had completed screens for tuberculosis, HIV, syphilis and hepatitis C. Tuberculosis screens were positive in 3.6 % of children, 5.5 % of adolescents were positive for chlamydia and <1 % of children were positive for HIV, syphilis or hepatitis C. Increasing age and number of visits were associated with completed tuberculosis, HIV, syphilis and hepatitis C screenings (p < 0.01); female adolescents with completed labs were more likely to be screened for gonorrhea and chlamydia than male adolescents. ConclusionsFew positive infectious disease screenings were identified in children evaluated in a dedicated foster care primary care clinic despite presence of risk factors. Multiple visits to a primary care foster care clinic may increase the likelihood of completed screenings. Targeted infectious disease screening based on age and local epidemiology may be less traumatizing but still clinically appropriate.

A tetravalent live attenuated dengue virus vaccine stimulates balanced immunity to multiple serotypes in humans

The four-dengue virus (DENV) serotypes infect several hundred million people annually. For the greatest safety and efficacy, tetravalent DENV vaccines are designed to stimulate balanced protective immunity to all four serotypes. However, this has been difficult to achieve. Clinical trials with a leading vaccine demonstrated that unbalanced replication and immunodominance of one vaccine component over others can lead to low efficacy and vaccine enhanced severe disease. The Laboratory of Infectious Diseases at the National Institutes of Health has developed a live attenuated tetravalent DENV vaccine (TV003), which is currently being tested in phase 3 clinical trials. Here we report, our study to determine if TV003 stimulate balanced and serotype-specific (TS) neutralizing antibody (nAb) responses to each serotype. Serum samples from twenty-one dengue-naive individuals participated under study protocol CIR287 (ClinicalTrials.gov NCT02021968) are analyzed 6 months after vaccination. Most subjects (76%) develop TS nAbs to 3 or 4 DENV serotypes, indicating immunity is induced by each vaccine component. Vaccine-induced TS nAbs map to epitopes known to be targets of nAbs in people infected with wild type DENVs. Following challenge with a partially attenuated strain of DENV2, all 21 subjects are protected from the efficacy endpoints. However, some vaccinated individuals develop post challenge nAb boost, while others mount post-challenge antibody responses that are consistent with sterilizing immunity. TV003 vaccine induced DENV2 TS nAbs are associated with sterilizing immunity. Our results indicate that nAbs to TS epitopes on each serotype may be a better correlate than total levels of nAbs currently used for guiding DENV vaccine development.

Development of a real-time Reverse-Transcription PCR for SARS CoV-2 on the Luminex ARIES® Platform

The University of Louisville Infectious Diseases Laboratory followed the US Food and Drug Administration (FDA) Emergency Use Authorization (EUA) guidance for developing a molecular diagnostic test for SARS CoV-2 to help address the novel coronavirus pandemic. As a Clinical Laboratory Improvement Amendment ‘88 (CLIA) certified, high-complexity clinical laboratory, the Infectious Diseases Laboratory chose to use the Luminex ARIES® platform to evaluate a laboratory developed test. This instrument was already familiar to the Infectious Diseases Laboratory and in use for molecular diagnostic testing for pathogens causing atypical pneumonia and two tick-borne pathogens. The FDA EUA guidance for molecular diagnostic tests recommended limit of detection studies, inclusivity and exclusivity (specificity) analysis, and validation with clinical samples to ensure the performance of the assay was acceptable for use as a molecular diagnostic tool. Data obtained from these experiments demonstrated acceptable performance per FDA guidance, as well as for CLIA requirements. Thus, the real-time Reverse Transcription PCR assay was implemented for diagnostic use on March 27, 2020 and was a great benefit to the local community in responding to the pandemic.

State-of-the-Art Data Management: Improving the Reproducibility, Consistency, and Traceability of Structural Biology and in Vitro Biochemical Experiments.

Efficient and comprehensive data management is an indispensable component of modern scientific research and requires effective tools for all but the most trivial experiments. The LabDB system developed and used in our laboratory was originally designed to track the progress of a structure determination pipeline in several large National Institutes of Health (NIH) projects. While initially designed for structural biology experiments, its modular nature makes it easily applied in laboratories of various sizes in many experimental fields. Over many years, LabDB has transformed into a sophisticated system integrating a range of biochemical, biophysical, and crystallographic experimental data, which harvests data both directly from laboratory instruments and through human input via a web interface. The core module of the system handles many types of universal laboratory management data, such as laboratory personnel, chemical inventories, storage locations, and custom stock solutions. LabDB also tracks various biochemical experiments, including spectrophotometric and fluorescent assays, thermal shift assays, isothermal titration calorimetry experiments, and more. LabDB has been used to manage data for experiments that resulted in over 1200 deposits to the Protein Data Bank (PDB); the system is currently used by the Center for Structural Genomics of Infectious Diseases (CSGID) and several large laboratories. This chapter also provides examples of data mining analyses and warnings about incomplete and inconsistent experimental data. These features, together with its capabilities for detailed tracking, analysis, and auditing of experimental data, make the described system uniquely suited to inspect potential sources of irreproducibility in life sciences research.

Bilateral Anterior Ischaemic Optic Neuropathy as First Manifestation of Henoch-Schönlein Purpura (IgA Vasculitis).

A 46-year-old man was referred to our department complaining of a bilateral progressive decrease in his visual acuity. Fundus examination revealed bilateral optic disc oedema, indicative of anterior ischaemic neuropathy (AION), and a macular star in the right eye. Laboratory analysis showed low haematocrit and haemoglobin, elevated creatinine, and increased erythrocyte segmentation rate and C-reactive protein level. Physical examination revealed the presence of purpuric rash on the trunk and the extremities. During the investigation we performed a complete laboratory and imaging examination for autoimmune collagen diseases, vasculitides and infectious diseases, which were all negative. Histologic findings of renal biopsy were compatible with IgA glomerulonephritis and thus Henoch-Schönlein purpura (HSP) diagnosis was established. The patient was treated with methylprednisolone and cyclophosphamide. Six months later, his renal function and his visual acuity had improved, and the rash had subsided. This is a rare case of AION in a patient with HSP.

Respiratory diseases in HIV-infected patients (review)

The review presents information on epidemiology, clinical, laboratory and radiological manifestations of infectious and malignant respiratory diseases in HIV-infected patients.The data on the most common infections (communityacquired pneumonia, tuberculosis, Pneumocystis jiroveci pneumonia) and malignant diseases (Kaposhi’s sarcoma, lymphomas) as well as less frequent diseases (CMV-infection, MAC-infection, fungal pneumonias) are presented.The article contains facts about the incidence of pulmonary diseases according to the severity of HIV-associated immunodeficiency. This review also presents current information on the diagnostic criteria, treatment regimens and secondary prophylaxis for HIV-associated respiratory infections and malignancies. The specialties of ART initiation in definite cases of opportunistic infections and ART assignment during the diagnostic period are highlighted.The growing number of HIV-infected patients and the wide spectrum of possible pulmonary diseases argue the need for optimal clinical approach to HIV-infected patients with respiratory lesions for various physicians (general practitioners, pulmonologists and phthisiatricians).

Letter from Singapore: The clinical and research response to COVID‐19

Letter from Singapore: The clinical and research response to COVID‐19

Infections after hematopoietic cell transplantation are not a burden for mortality at a limited-resource center in a developing country.

Hematopoietic cell transplantation (HCT) outcomes, including infectious complications, change between centers and countries. Thus, the aim of this study was to report the incidence of infections and isolated pathogens among recipients of HCT and the association with mortality at a tertiary referral center in Mexico. Two hundred and eighty-two patients undergoing autologous or allogeneic HCT between January 2005 and December 2018 at the National Institute of Medical Sciences and Nutricion Salvador Zubiran were included. In autologous HCT (n=176), within the preengraftment and the early postengraftment, 130 (74%) and 31 (18%) recipients presented infections, respectively. Within the preengraftment, the early postengraftment, and the late postengraftment, 81 (76%), 34 (33%), and 58 (60%) allogeneic HCT recipients presented infections, respectively. Non-relapse mortality (NRM) as a result of infections occurred in 1 (0.6%) and 5 (5%) autologous and allogeneic HCT recipients, respectively. Our results demonstrated that despite our limited resources, infections were not a significant burden for NRM among HCT recipients. More importantly, the isolation rates were higher than international studies, which could be explained by the existence of a specialized infectious diseases department and laboratory, which we consider key elements for the establishment of an HCT program worldwide.

Etiologies of severe acute respiratory infection (SARI) and misdiagnosis of influenza in Indonesia, 2013-2016.

BackgroundSevere acute respiratory infection (SARI) accounts for a large burden of illness in Indonesia. However, epidemiology of SARI in tertiary hospitals in Indonesia is unknown. This study sought to assess the burden, clinical characteristics, and etiologies of SARI and concordance of clinical diagnosis with confirmed etiology.MethodsData and samples were collected from subjects presenting with SARI as part of the acute febrile Illness requiring hospitalization study (AFIRE). In tertiary hospitals, clinical diagnosis was ascertained from chart review. Samples were analyzed to determine the “true” etiology of SARI at hospitals and Indonesia Research Partnership on Infectious Diseases (INA‐RESPOND) laboratory. Distribution and characteristics of SARI by true etiology and accuracy of clinical diagnosis were assessed.ResultsFour hundred and twenty of 1464 AFIRE subjects presented with SARI; etiology was identified in 242 (57.6%), including 121 (28.8%) viruses and bacteria associated with systemic infections, 70 (16.7%) respiratory bacteria and viruses other than influenza virus, and 51 (12.1%) influenza virus cases. None of these influenza patients were accurately diagnosed as having influenza during hospitalization.ConclusionsInfluenza was misdiagnosed among all patients presenting with SARI to Indonesian tertiary hospitals in the AFIRE study. Diagnostic approaches and empiric management should be guided by known epidemiology. Public health strategies to address the high burden of influenza should include broad implementation of SARI screening, vaccination programs, clinician education and awareness campaigns, improved diagnostic capacity, and support for effective point‐of‐care tests.

Assessment of Antiplasmodial and Antioxidant Activities, Total Phenolics and Flavonoids Content, and Toxicological Profile of Cola millenii K. shum (Malvaceae)

Aims: This study was designed to assess antiplasmodial and antioxidant activities in relation to phytochemical contents and toxicological profile of crude extracts of Cola millenii leaves.&#x0D; Place and Duration of Study: The study was carried out from April 2015 to November 2018 at University of Abomey-Calavi, Laboratory of Biochemistry and Bioactives Natural Products and Laboratory of Infectious Vector-borne Diseases.&#x0D; Methodology: Phytochemical screening of aqueous and ethanolic crude extracts was performed using standard methods. Estimation of total phenolics contains (TPC) and total flavonoids contains (TFC) was done using colorimetric methods. Antioxidant activity was assessed in vitro by 2,2-diphenyl-1-picrylhydrazyl (DPPH) Radical-Scavenging, Reducing power, superoxide radical scavenging and Hydrogen peroxide scavenging assay. Antiplasmodial activity was investigated in vitro using both field isolate and laboratory 3D7 strains of P. falciparum using Plasmodium lactate dehydrogenase (pLDH) assay. Hemoglobin release was measured spectrophotometrically to assess hemolytic power. Acute oral toxicity of extracts was evaluated on rats according to OECD 423 guideline.&#x0D; Results: Flavonoids, anthraquinones, coumarins, and triterpenes were recorded in both extracts while tannins were recorded only in ethanolic extract. High TPC and TFC were recorded with ethanolic extract with significate difference (P &lt;0.01) when compared to aqueous extract. Ethanolic extract exhibited the highest parasite growth inhibition against both field isolate (41.62±1.78%) and 3D7 (45.89±1.66%) strains at 100 µg/mL. Both extracts exhibited strong antioxidant activity according to different methods used. Toxicological profile revealed no hemolytic effect on human red blood cells as well as acute toxicity signs in rats.&#x0D; Conclusion: This study demonstrated strong antioxidant and moderate antiplasmodial activities of C. millenii extracts without toxicity effect on rats and human erythrocytes. It would play an important role in malaria and oxidative damage control.

Multiomics Evaluation of Gastrointestinal and Other Clinical Characteristics of COVID-19

Since December 2019, coronavirus disease 2019 (COVID-19) outbreak caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has produced a worldwide panic. Beyond the principal human-to-human transmission method by droplet and contact, there is still limited knowledge about possible alternate transmission methods to guide clinical care. Recent clinical studies have observed digestive symptoms in patients with COVID-19,1 possibly because of the enrichment and infection of SARS-CoV-2 in the gastrointestinal tract, mediated by virus receptor of angiotensin converting enzyme 2 (ACE2),2 which suggests the potential for a fecal-oral route of SARS-CoV-2 transmission.3,4 However, there is still a large gap in the biological knowledge of COVID-19. In this study, via a bulk-to-cell strategy focusing on ACE2, we performed an integrated omics analysis at the genome, transcriptome, and proteome levels in bulk tissues and single cells across species to decipher the potential routes for SARS-CoV-2 infection in depth.

Gastrointestinal organoid technology advances studies of enteric virus biology.

The historical lack of models recapitulating the complexity of the human intestinal epithelium has hindered studies into many aspects of human enteric virus biology. Immortalized and transformed cell lines are typically limited by the presence of only one cell type, whereas susceptibility in animal models often requires infection routes that differ from humans or necessitates modification of immune system components [1, 2]. In addition, comparing animals from different species remains a confounding factor when trying to infer how findings may apply to human health. Thus, the development of human gastrointestinal organoids to study virus–host interactions marks a significant advance. They provide a physiologically relevant ex vivo platform in which human enteric microorganisms can be studied interacting with the human intestinal epithelium. Furthermore, their intermediate complexity, falling between cell lines and animal models, adds an additional tool to better understand these viruses. Here, we highlight how gastrointestinal organoids have provided new insights into the biology of human enteric viruses and the potential of this technology for advancing the field. The different flavors of gastrointestinal organoids Gastrointestinal organoids are three-dimensional (3D) structures derived from primary tissues (i.e., patient biopsy) containing intestinal stem/progenitor cells or from human pluripotent stem cells (hPSCs) [3]. They contain multiple intestinal epithelial cell types that perform critical functions that are also observed in the human intestine (e.g., absorption, barrier function, differentiation). Induced pluripotent stem cell–derived human intestinal organoids (HIOs) most closely resemble the human fetal intestine [4] and may encompass an epithelium alone [5] or also contain a mesenchyme [6]. However, most infectious disease laboratories work with human intestinal enteroids (HIEs), patient-derived, 3D epithelium-only structures that can be transitioned to a 2D monolayer on plates or transwells. HIEs maintain the physiological and genetic characteristics of their sources for long periods [7–10]. They can be differentiated from the crypt-like state into villus-like state by withdrawing growth factors required to maintain stem cells (i.e., WNT3A) from the culture media [11–13]. Cellular differentiation of specific cell lineages can be further achieved by pharmacologic or genetic means. For example, secretory cells are enriched following dibenzazepine (DBZ) treatment to block NOTCH signaling [12] and enteroendocrine cells by overexpression of NEUROGENIN-3 [14]. In addition, receptor activator of NF-κB ligand (RANKL) treatment of HIE with and without tumor necrosis factor alpha (TNF-α) addition has been used to drive microfold cell development [15, 16]. However, HIE and HIO do not completely mimic the intestinal epithelium in vivo. For example, they do not entirely reproduce the cellularity observed in vivo as few Paneth cells and no Tuft cells are present [17]. Furthermore, the lack of villi and genetic or pharmacologic skewing of differentiation may also affect the proportionality of individual cell lineages.

Bilateral anterior ischaemic optic neuropathy with macular star as first manifestation of Henoch‐Schönlein purpura (IgA vasculitis)

AbstractPurposeTo report a case of bilateral anterior ischaemic optic neuropathy with macular star in a patient with Henoch–Schönlein purpura (HSP).MethodsA 46‐year‐old man was referred to our clinic complaining of progressive decrease in visual acuity of the right eye (OD) since the last 10 days, and since 3 days in his left eye (OS). His visual acuity was ‘hand motion’ OD and 4/10 OS. Anterior segment examination was insignificant, while fundus examination revealed optic disc edema, and bilateral macular star. A relative afferent pupillary defect was present in his right eye with normal pupillary reflexes in the left eye. Laboratory analysis showed: Hematocrit: 22.1%, ESR: 140 mm/hr, CRP: 64.80 mg/dl, Creatinine: 4.4 mg/dl. Physical examination revealed presence of erythematous rash on the trunk and the extremities. The patient was admitted to the Nephrology department for treatment of his azotemia.ResultsDuring his hospital course we performed complete laboratory and imaging examinations for autoimmune diseases, vasculitis and infectious diseases, which were negative. Histologic findings of renal biopsy were compatible with IgA glomerulonephritis and thus HSP was diagnosed. The patient received oral methylprednisolone and intravenous cyclophosphamide therapy. Six months later his renal function had improved, the rash had subsided and his visual acuity was ‘counting fingers’ OD and 10/10 OS. Standard automated perimetry showed peripheral visual field depression bilaterally.ConclusionsHenoch–Schönlein purpura (HSP) is a small vessel vasculitis which is more common in children. Based on literature, ophthalmic manifestations of HSP usually involve the anterior segment and less frequently the posterior segment. Systemic vasculitis should be included in the differential diagnosis of bilateral optic disc edema.

Qatar National Paediatric Sepsis Program: Where are we?

Background: Sepsis and septic shock are medical emergencies and necessitate early and timely recognition and intervention. Failure of early recognition can lead to significant deterioration and may unfortunately culminate in death.1,2 Young people are considered at high risk of sepsis.3,4 Despite hundreds of trials and a multitude of approaches, an effective and efficient sepsis-cure agent does not exist. Most research into sepsis management has ended with non-conclusive and sometimes confusing results. Current evidence recommends a bundle of simple interventions to be accomplished as soon as possible and preferably within the first hour of sepsis recognition.2,5 A number of international initiatives aim at reducing sepsis mortality.2 Recently the World Health Organization (WHO) urged governments to set national mechanisms to tackle sepsis. A nation-wide sepsis program was developed to improve sepsis care for people in Qatar. A parallel National Paediatric Sepsis Program was developed to provide appropriate guidelines, education, a unified national care pathway, and to increase awareness of paediatric sepsis. Here we discuss the design and outcome of the Qatar paediatric sepsis program to date.Methods: The program aims for early sepsis recognition and 95% compliance with the Sepsis Care Bundle by the end of 2019. To achieve this target a multi-faceted approach to paediatric sepsis care across all public healthcare sectors in Qatar was adopted. This includes Sidra Medicine and Hamad Medical Corporation and its Paediatric Emergency Centres.An overarching system-wide sepsis committee was established and included major stakeholders within emergency medicine, critical care, infection prevention and control, and infectious disease and clinical laboratory. A paediatric multidisciplinary sepsis committee was established in 2017 and the National Paediatric Sepsis program was developed.International evidence-based Institute for Health Improvement (IHI) methodology was adopted for program development. Major areas of the program were dedicated to the formulation of clinical practice guidelines, standardised care pathway, standardised EMR order set for all clinical areas, and ongoing education and awareness for healthcare providers at all facilities.Sepsis simulation sessions were conducted to fill knowledge gaps and an improvement ramp module was included based on the PDSA (Plan-Do-Study-Act) strategy. A number of other PDSA initiatives were undertaken and included the following: Establish the paediatric sepsis clinical pathway and guideline (Figure 1); introduce the sepsis watchers’ concept to daily safety huddle; provide a standardized paediatric sepsis diagnostic kit; create unified paediatric sepsis antibiotics kits for all units with a safe first dose preparation protocol; develop and implement an e-learning module with education materials and paediatric sepsis order set in the electronic system.Ongoing data collection and performance evaluation for sustainability and dissemination of information demonstrated the following:1. Paediatric sepsis incidence varies per facility and over time. Between 30 to 100 cases/month (Figure 2).2. Recognition: Percentage of Clinical Review, Rapid Response Team (RRT) activation, and sepsis alerts that were appropriately escalated is 91%.3. Order set use: 26% initiating the well-established electronic paediatric sepsis golden-hour order set.4. 42% bundle compliance (Figure 3).5. IV antibiotics within 60 minutes of time zero showed 81% compliance. Conclusion: Current literature suggests that systemic and supervised implementation of an evidence-based pathway for suspected and confirmed paediatric sepsis saves lives. Our data demonstrated poor bundle compliance but significant improvement is seen in the areas of early recognition and antibiotic administration within one hour. Education and awareness are key to improve performance.

Implementation of an Algorithmic Approach to Molecular Infectious Disease Laboratory Testing on Cerebrospinal Fluid Samples

Abstract Rapid turnaround time for infectious disease laboratory testing is critical for the encephalitic/meningitic patient. Clinicians and emergency medicine providers often will bundle cerebrospinal fluid (CSF) studies (cell counts, glucose, protein) with molecular infectious disease laboratory (MIDL) assays, up front, to expedite clinical decision making about antimicrobial therapy and hospital admission. Many patients, however, ultimately do not have CSF pleocytosis and, predictably, have negative MIDL results. We quantified patients who had MIDL testing ordered, without concurrent CSF pleocytosis, to justify a mandated algorithm for CSF MIDL testing. Four months of MIDL CSF results were reviewed (viral monoplex PCRs); 274 patients were identified. Forty-seven percent of patients (128/274) were neonates (&lt;2 months old) and thus may not have had the immunologic capacity to mount a pleocytosis to CNS infection. Of the remaining 53% of patients (&gt;2 months old), who would have been expected to generate a CSF pleocytosis, 55% (81/146) had one or more MIDL studies ordered without CSF pleocytosis; 3.4% of these patients (5/146) did not have CSF cell counts ordered. Among all patients, 7.7% (21/274) had positive MIDL results (1 HSV1; 2 HSV2; 18 enterovirus). Seventeen patients had CSF pleocytosis; the 4 patients without CSF pleocytosis were neonates (3) or had no cell counts ordered (1). A ratio of 700 RBC to 1 WBC was used to approximate the expected number of CSF WBC in bloody taps (10%; 28/274). We show that over half of patients &gt;2 months old had MIDL studies ordered without CSF pleocytosis; all patients with positive MIDL results had CSF pleocytosis or were neonates or had no cell counts ordered. An algorithm will be implemented in which all neonates with concern for CNS infection will have CSF MIDL tests ordered; among nonneonates, only those with documented CSF pleocytosis may have MIDL assays ordered (with rare exceptions; eg, leukopenia).

Anti-Chlamydia abortus and Anti-Mycobacterium avium subsp. paratuberculosis Antibodies in Buffaloes in the State of Pernambuco, Brazil

Background: The occurrence of economic losses in buffaloes may be related to reproductive problems such as chlamydiosis caused by the bacteria Chlamydia abortus considered as a zoonotic agent; and digestive problems highlighting the infection by Mycobacterium avium subsp. paratuberculosis (Map), responsible for paratuberculosis. There is a little information about these diseases in buffaloes, therefore the aim of this study was to determine the occurrence of anti-Chlamydia abortus and anti- Mycobacterium avium subsp. paratuberculosis (Map) antibodies in water buffaloes in the state of Pernambuco, Brazil.Materials, Methods &amp; Results: The 262 bubaline sera belonging to the serum bank of the Infectious Diseases Laboratory (LIDIC) of the Federal Rural University of Pernambuco (UFRPE) were analyzed. The samples were from nine properties distributed in the municipalities of Agreste and Zona da Mata of the state of Pernambuco. For the detection of anti-Chlamydia abortus and anti-Map antibodies was used a technique of Enzyme immunoabsorption Assay (ELISA) of the IDEXX® by following the manufacturer's instructions. Regarding the detection of anti-Chlamydia abortus antibodies, it was observed that 47.70% (125/262) of the samples were positive. All properties showed at least one positive animal for the investigation of anti-Chlamydia abortus antibodies. It was also verified the occurrence of 7.25% of suspected animals for the investigation of anti-C. abortus antibodies. No positives animals were observed for Map in the bubaline analyzed.Discussion: The occurrence of anti-C. abortus antibodies in buffaloes in the region can be explained by the fact that properties with buffalo breeding has sanitary management that allows the contact between animals, thus increasing the risk of transmission of the agent. In addition, the variation found from 35% to 68.75% may be associated with divergences to the type of management and breeding system used in each property. The percentage of suspected animals may suggest that the number of positive animals is higher or that there were non-specific reactions with other species of Chlamydia, but it is not possible to determine the seroconversion without the accomplishment of paired serology. Because it is a zoonosis, C. abortus may be a risk to the health of the population involved, since the transmission of the bacteria to humans can occur by contact with secretions and excretions of these animals. No positive animals were found for the occurrence of anti-Map antibodies, however, there are reports of the infection identifying animals with clinical signs or properties with a history of the disease, and this may be related to the differences in each property in relation to the history and sanitary management. Moreover, the ELISA test may not be sensitive to the agent depending on the phase of the infection, because if the response is predominantly cellular, the number of antibodies is diminished, making the serological diagnosis difficult. The use of more sensitive tests for the bubaline species can also favor the diagnosis of the infection. Although the occurrence of anti-Map antibodies in the analyzed samples was not detected, it is important to carry out routine epidemiological studies, since the disease has already been registered in water buffaloes in the state of Pernambuco. Considering the occurrence of anti-Chlamydia abortus antibodies it is suggested to conduct studies in order to isolate the agent and verify its real importance in the reproductive disorders of the bubaline species, since this agent causes reproductive losses and has a zoonotic character.

The Cano-eMLST Program: An Approach for the Calculation of Canonical Extended Multi-Locus Sequence Typing, Making Comparison of Genetic Differences Among Bunches of Bacterial Strains.

Extended multi-locus sequence typing (eMLST) methods have become popular in the field of genomic epidemiology. Before eMLST methods can be applied in epidemiological investigations, the selection of a suitable scheme is critical. The core genome scheme (cgMLST) has become the most popular eMLST approach for strain typing in the epidemiological domain. In addition to strain typing, many public health researchers and clinical microbiologists wish to investigate which genes cause genetic differences between compared strains. Therefore, a tool that can be used to extract canonical genes with an eMLST scheme would be particularly useful. In this study, we present cano-eMLST, a well-designed program that applies a feature-selection methodology to create a canonical locus combination with discriminatory power by traversing a genetic relatedness tree based on a user-selected scheme. The cano-eMLST program is provided mainly to help infectious disease laboratory researchers identify potential factors related to bacterial pathogenesis. The core program (tree-traversing approach) of cano-eMLST is implemented in Perl and Python. All the necessary dependencies and environmental settings are provided in the encapsulated version (VirtualBox or VMware) and self-installation version (all use source code and libraries).