Determination of resistance status to amitraz in the cattle tick Rhipicephalus (Boophilus) microplus from Luzon, Philippines, through bioassay and molecular analysis.

Tick infestations and tick-borne diseases are among the leading causes of economic losses in the cattle industry worldwide. Amitraz is one of the most commonly used chemical acaricides against the cattle tick Rhipicephalus (Boophilus) microplus. Resistance to amitraz has been reported in many countries but not in the Philippines. This study aimed to determine whether cattle ticks from provinces in northern and southern Luzon, Philippines, are resistant to amitraz. Engorged or nearly engorged female ticks were collected from 21 farms and allowed to lay eggs to produce larvae. Larval packet test (LPT) was performed using three concentrations of amitraz, as well as a negative control (diluent). Reverse-transcription polymerase chain reaction (RT-PCR) was also performed to amplify a fragment of the octopamine/tyramine receptor gene and was subjected to sequence analysis by multiple nucleotide and amino acid sequence alignments together with reference strain, amitraz-susceptible strain, and amitraz-resistant strain reported in other countries. LPT results suggest the absence of resistance in all of the tested populations. Interestingly, analysis of the octopamine/tyramine receptor amino acid sequence revealed four out of 14 larval pools having substitutions similar to that of the reported amitraz-resistant strains. Although no apparent resistance was observed in this study, prudent use of amitraz should be practiced as the development of resistance is still likely to occur in the future.

A Comparative Analysis of Pulmonary and Critical Care Medicine Guideline Development Methodologies.

RATIONALE: The Institute of Medicine (IOM) standards for guideline development have had unintended negative consequences. A more efficient approach is desirable. OBJECTIVES: To determine whether a modified Delphi process early during guideline development discriminates recommendations that should be informed by a systematic review from those that can be based upon expert consensus. METHODS: The same questions addressed by IOM-compliant pulmonary or critical care guidelines were addressed by expert panels using a modified Delphi process, termed the Convergence of Opinion on Recommendations and Evidence (CORE) process. The resulting recommendations were compared. Concordance of the course of action, strength of recommendation, and quality of evidence, as well as the duration of recommendation development, were measured. MEASUREMENTS AND MAIN RESULTS: When 50% agreement was required to make a recommendation, all questions yielded recommendations, and the recommended courses of action were 89.6% concordant. When 70% agreement was required, 17.9% of questions did not yield recommendations, but for those that did, the recommended courses of action were 98.2% concordant. The time to completion was shorter for the CORE process (median, 19.3 vs. 1,309.0 d; P = 0.0002). CONCLUSIONS: We propose the CORE process as an early step in guideline creation. Questions for which 70% agreement on a recommendation cannot be achieved should go through an IOM-compliant process; however, questions for which 70% agreement on a recommendation can be achieved can be accepted, avoiding a lengthy systematic review.

Acute-Phase Deaths from Murine Polymicrobial Sepsis Are Characterized by Innate Immune Suppression Rather Than Exhaustion.

Sepsis, a leading cause of death in the United States, has poorly understood mechanisms of mortality. To address this, our model of cecal ligation and puncture (CLP) induced sepsis stratifies mice as predicted to Live (Live-P) or Die (Die-P) based on plasma IL-6. Six hours post-CLP, both Live-P and Die-P groups have equivalent peritoneal bacterial colony forming units and recruitment of phagocytes. By 24 h, however, Die-P mice have increased bacterial burden, despite increased neutrophil recruitment, suggesting Die-P phagocytes have impaired bacterial killing. Peritoneal cells were used to study multiple bactericidal processes: bacterial killing, reactive oxygen species (ROS) generation, and phagocytosis. Total phagocytosis and intraphagosomal processes were determined with triple-labeled Escherichia coli, covalently labeled with ROS- and pH-sensitive probes, and an ROS/pH-insensitive probe for normalization. Although similar proportions of Live-P and Die-P phagocytes responded to exogenous stimuli, Die-P phagocytes showed marked deficits in all parameters measured, thus suggesting immunosuppression rather than exhaustion. This contradicts the prevailing sepsis paradigm that acute-phase sepsis deaths (<5 d) result from excessive inflammation, whereas chronic-phase deaths (>5 d) are characterized by insufficient inflammation and immunosuppression. These data suggest that suppression of cellular innate immunity in sepsis occurs within the first 6 h.

The health-system benefits and cost-effectiveness of using Mycobacterium tuberculosis direct nucleic acid amplification testing to diagnose tuberculosis disease in the United States.

BACKGROUND: The utility of Mycobacterium tuberculosis direct nucleic acid amplification testing (MTD) for pulmonary tuberculosis disease diagnosis in the United States has not been well described. METHODS: We analyzed a retrospective cohort of reported patients with suspected active pulmonary tuberculosis in 2008-2010 from Georgia, Hawaii, Maryland, and Massachusetts to assess MTD use, effectiveness, health-system benefits, and cost-effectiveness. RESULTS: Among 2140 patients in whom pulmonary tuberculosis was suspected, 799 (37%) were M. tuberculosis-culture-positive. Eighty percent (680/848) of patients having acid-fast-bacilli-smear-positive specimens had MTD performed; MTD positive-predictive value (PPV) was 98% and negative-predictive value (NPV) was 94%. Nineteen percent (240/1292) of patients having smear-negative specimens had MTD; MTD PPV was 90% and NPV was 88%. Among patients suspected of tuberculosis but not having MTD, smear PPV for lab-confirmed tuberculosis was 77% and NPV 78%. Compared with no MTD, MTD significantly decreased time to diagnosis in patients with smear-positive/MTD-positive specimens, decreased respiratory isolation for patients having smear-positive/MTD-negative/culture-negative specimens, decreased outpatient days of unnecessary tuberculosis medications, and reduced resources expended on contact investigation. While MTD generally cost more than no MTD, incremental cost savings occurred in patients with human immunodeficiency virus (HIV) or homelessness to diagnose or to exclude tuberculosis, and in patients with substance abuse having smear-negative specimens to exclude tuberculosis. CONCLUSIONS: MTD improved diagnostic accuracy and timeliness and reduced unnecessary respiratory isolation, treatment, and contact investigations. It was cost saving in patients with HIV, homelessness, or substance abuse, but not in others.

Factors Associated with Development of Tuberculosis Disease Among Refugees, Massachusetts, 2008-2018.

Refugees and immigrants undergo tuberculosis screening prior to arrival in the United States. CDC Technical Instructions for screening changed in 2007. Our goal was to quantify TB disease in refugees after 2007 and identify risks for disease. Massachusetts refugee and tuberculosis databases were matched to identify refugees who arrived 2008-2017 and were diagnosed with tuberculosis infection or disease 2008-2018. Factors associated with disease were analyzed in SAS. Of 19,583 refugees, 4706 were diagnosed with infection at arrival and 60 with disease during the observation period. Lack of treatment for infection was strongly associated (OR = 26.5, p = 0.0001) with diagnosis of disease; in a multivariate logistic regression model, positive screening test (AOR = 12.5, p = 0.0001), class B1 status (AOR = 4.0, p = 0.0004), and < 2 years since arrival (AOR = 60.0, p = 0.0001) were associated with disease. Providers should continue screening new arrivals, providing accessible services, and treating infection to further reduce tuberculosis morbidity and mortality.

Nonclinical selection criteria for maximizing yield of nucleic acid amplification tests in tuberculosis diagnosis.

In spite of the excellent performance of rapid tuberculosis (TB) nucleic acid amplification (NAA) tests and the clear benefits of immediate diagnosis of TB disease, NAA tests frequently are not used in the diagnosis of pulmonary TB cases, particularly TB cases with smear-negative sputa. Public health laboratories primarily perform TB NAA tests only on a targeted subset of specimens, usually including those that are smear positive and those for which a clinician has specifically requested NAA testing. As an alternative to targeted testing, some laboratories use TB NAA tests universally for all respiratory specimens, though this practice can be prohibitively costly and can be associated with an increased frequency of false-positive results due to testing of lower-risk patients. We propose a strategy for identifying individuals for NAA testing on the basis of nonclinical risk criteria that are routinely provided on the test requisition form, such as type of health care facility from which the specimen is received and patient age group. Use of this strategy at the Massachusetts Department of Public Health Laboratory would allow for NAA test identification of approximately 54 (74%) of 72 culture-positive pulmonary TB cases over a 1-year period while requiring NAA testing for only 933 (17%) of 5,469 individuals submitting respiratory specimens. We demonstrate that use of nonclinical NAA test selection criteria is an effective strategy for maximizing the number of TB cases that can be rapidly identified while minimizing the number of specimens that must be tested.

Expanding Latent Tuberculosis Infection Testing and Treatment in Massachusetts Primary Care Clinics via the ECHO Model.

INTRODUCTION/OBJECTIVES: In the US, reactivation of latent tuberculosis infection (LTBI) accounts for 80% of new cases. In 2016, the US Preventive Services Task Force provided a new recommendation that primary care providers (PCPs) should conduct LTBI screening, whereas in the past, LTBI cases were evaluated and treated by specialty providers. This shift in care revealed knowledge gaps surrounding LTBI treatment among PCPs. This study assessed changes in PCPs' confidence for performing key aspects of LTBI care before and after participation in an LTBI Extension for Community Healthcare Outcomes (ECHO) course. METHODS: The ECHO Model™ is an evidence-based telementoring intervention. Participants were primary care team members from clinics throughout Massachusetts who voluntarily enrolled in the ECHO course. In this mixed-methods evaluation, primary outcomes were PCP self-reported confidence changes by pre- and post-course surveys and post-course semi-structured interviews. RESULTS: Twenty PCPs (43% of registered PCPs) attended at least 3 of the 6 sessions and 24 PCPs (31% of registered PCPs) completed at least one survey. Confidence increased in selecting a test (P = .004), interpreting tuberculosis infection test results (P = .03), and selecting a treatment regimen (P = .004). Qualitative interviews with 3 PCPs revealed practice changes including switching to interferon gamma release assays for testing and using rifampin for treatment. CONCLUSIONS: Use of the ECHO model to train PCPs in LTBI management is feasible and efficacious. For continuing medical education, ECHO courses can be leveraged to reduce health disparities in settings where PCPs' lack of familiarity about a treatment topic contributes to poor health outcomes.

Molecular Detection and Characterization of Babesia and Theileria in Cattle and Water Buffaloes from Southern Luzon, Philippines.

Babesia and Theileria are tick-borne protozoan parasites that can cause significant economic losses in the cattle industry. This study aimed to contribute to the limited epidemiological data on Theileria orientalis as well as Babesia bigemina and B. bovis in large ruminants in the Philippines. Blood samples of 412 cattle and 108 water buffalo collected from four provinces in Southern Luzon, Philippines, were initially tested for the 18S rRNA gene of piroplasms through nested PCR. Positive samples were further subjected to species-specific PCR. The 18s rRNA of piroplasms was detected in 123 (29.9%) cattle and three (2.8%) water buffaloes. Theileria orientalis was found to be the most common piroplasm in cattle with a detection rate of 17.5%, followed by Babesia bovis and B. bigemina. Co-infections were also observed. Two water buffaloes were found infected with B. bovis, while one was positive for B. bigemina. The phylogenetic tree for B. bovis showed clustering of the isolates in two clades together with isolates from other countries, and a third separate clade. Meanwhile, the T. orientalis isolates in this study were distributed in three clades together with reported isolates from other countries. This study confirms the presence of T. orientalis in the Philippines and reports the genetic diversity of B. bovis and T. orientalis.

The need for performance measures on testing for latent tuberculosis infection in primary care.

BACKGROUND: The millions of people living in the United States with latent tuberculosis infection (LTBI) represent a reservoir of potentially active tuberculosis (TB) disease. When LTBI is left to activate, the consequences may include intense suffering, permanent disability, and high economic costs for patients, their caretakers, and society at large as TB spreads. The introduction of performance measures would improve accountability for quality of care and to reduce disparities, especially if the measures are group-targeted. PERFORMANCE MEASURES PROPOSAL: One National Quality Forum-endorsed measure (#0408) calculates the rate of TB screening in persons with HIV. Using the measure as a model, a set of performance measures is proposed. Denominators will include all persons in a given high-risk category, and numerators will include those persons from the denominators with LTBI test results. National guidelines informed appropriate exclusions. IMPLEMENTATION CHALLENGES AND SOLUTIONS: Challenges to implementation include lack of TB knowledge among primary care providers, potential for overwhelming already burdened schedules, and stigma associated with TB. However, the new measures, along with publication of educational resources, would raise clinicians' awareness. Short checklists and electronic supports would minimize time pressures. The routinization of screening would help reduce stigma. Finally, new federal funding and political will for electronic health records would facilitate data collection and impact assessment. CONCLUSIONS: TB sits at the crossroads of health and economic inequity and is a huge public health problem. The proposed performance measures will address a neglected secondary prevention opportunity and will be consistent with national priorities and health reform.

Initial cytoplasmic and phagosomal consequences of human neutrophil exposure to Staphylococcus epidermidis.

Microorganisms are recognized by specific phagocyte surface receptors. Liganded receptors then signal a series of events leading to phagocytosis and destruction of the organism by oxidative, lytic, and associated processes. Some organisms, such as Mycobacterium tuberculosis (Mtb), Cryptococcus neoformans (Cf), and others, evade such destruction, surviving and sometimes multiplying within the phagosome to later cause disease. To study such evasion, we developed protocols which permit simultaneous kinetic measurement of early cytoplasmic signaling and of phagosomal pH (pH(p)) and oxidative burst, on a cell-by-cell basis, of polymorphonuclear (PMN) leukocytes exposed to fluorescently labeled, nonpathogenic Staphylococcus epidermidis (Se). The availability of a new, highly sensitive pH probe, pHrodo, permits observation of increasing pH(p). Simultaneous labeling of the organism, applicable to any phagocyte target, with a probe insensitive to pH and oxidative species, such as AlexaFluor350, permits distinction between binding and functional responses to it by ratioing fluorescences. Addition of an extracellular-specific quencher (Trypan blue) permits distinction between bound and phagosome-enclosed targets, so that conditions within the closed phagosome can be studied. We found that opsonization is required for functional activation of PMN by Se, that the organism causes early alkalinization of the phagosome (in contrast to Cf which hyperacidifies it), and that extracellular Ca(2+) is not required for cytoplasmic Ca(2+) signaling but contributes markedly to binding of Se to PMN and to ensuant bactericidal functions. These findings lead to a new approach to the study of select organisms, like Cf and Mtb, which evade killing by manipulating the phagosomal environment.

Substitution of moxifloxacin for isoniazid during intensive phase treatment of pulmonary tuberculosis.

RATIONALE: Moxifloxacin has potent activity against Mycobacterium tuberculosis in vitro and in a mouse model of antituberculosis (TB) chemotherapy, but data regarding its activity in humans are limited. OBJECTIVES: Our objective was to compare the antimicrobial activity and safety of moxifloxacin versus isoniazid during the first 8 weeks of combination therapy for pulmonary TB. METHODS: Adults with sputum smear-positive pulmonary TB were randomly assigned to receive either moxifloxacin 400 mg plus isoniazid placebo, or isoniazid 300 mg plus moxifloxacin placebo, administered 5 days/week for 8 weeks, in addition to rifampin, pyrazinamide, and ethambutol. All doses were directly observed. Sputum was collected for culture every 2 weeks. The primary outcome was negative sputum culture at completion of 8 weeks of treatment. MEASUREMENTS AND MAIN RESULTS: Of 433 participants enrolled, 328 were eligible for the primary efficacy analysis. Of these, 35 (11%) were HIV positive, 248 (76%) had cavitation on baseline chest radiograph, and 213 (65%) were enrolled at African sites. Negative cultures at Week 8 were observed in 90/164 (54.9%) participants in the isoniazid arm, and 99/164 (60.4%) in the moxifloxacin arm (P = 0.37). In multivariate analysis, cavitation and enrollment at an African site were associated with lower likelihood of Week-8 culture negativity. The proportion of participants who discontinued assigned treatment was 31/214 (14.5%) for the moxifloxacin group versus 22/205 (10.7%) for the isoniazid group (RR, 1.35; 95% CI, 0.81, 2.25). CONCLUSIONS: Substitution of moxifloxacin for isoniazid resulted in a small but statistically nonsignificant increase in Week-8 culture negativity.

E-cigarette, or vaping, product use-associated lung injury: a review.

BACKGROUND: E-cigarette, or Vaping, Product Use-Associated Lung Injury (EVALI) is a disease entity related to the use of battery-operated or superheating devices that create an aerosolized form of nicotine and tetrahydrocannabinol (THC) and/or other substances for inhalation. METHODS: We performed a literature review to document epidemiology, pathogenesis and risk factors, diagnosis, clinical presentation, evaluation and management of EVALI. RESULTS: In the summer of 2019, an outbreak of EVALI cases brought this disease entity into the national spotlight. Since being recognized as a serious pulmonary disease with public health implications, more than 2600 cases have been reported to CDC with 68 deaths as of February 2020. The pathophysiology of EVALI remains unknown. Substances such as Vitamin E acetate have been implicated as a possible causes of lung injury. The CDC has established case definitions of "confirmed EVALI" cases to help guide identification of the disease and assist in surveillance. While clinical judgement by healthcare providers is imperative in the identification of EVALI cases, the heterogeneous presentations of EVALI make this difficult as well. Ultimately most investigative studies should be aimed at ruling out other disease processes that can present similarly. Treatment is centered around removing the offending substance and providing supportive care. CONCLUSIONS: EVALI is a serious pulmonary disease with public health implications. Diagnosis requires a high degree of suspicion to diagnose and exclusion of other possible causes of lung disease. It may be beneficial to involve a pulmonary specialist early in the management of this disease which is generally supportive care.

Increasing Drug Resistance Among Persons With Tuberculosis in Massachusetts, 2009-2018.

We examined Massachusetts tuberculosis surveillance data from to 2009 to 2018. Of 1533 culture-confirmed cases, 190 (12.4%) demonstrated resistance to isoniazid including 32 (2.1%) with rifampin resistance. In multivariable analysis, isoniazid resistance increased significantly over time (per-year odds ratio = 1.07, 95% confidence interval = 1.01-1.13, P = .018) and was associated with younger age, foreign birth, and prior tuberculosis treatment.

Prospective comparison of the tuberculin skin test and 2 whole-blood interferon-gamma release assays in persons with suspected tuberculosis.

BACKGROUND: Interferon-gamma release assays (IGRAs) are attractive alternatives to the tuberculin skin test (TST) for detecting Mycobacterium tuberculosis infection. However, the inability to definitively confirm the presence of most M. tuberculosis infections hampers assessment of IGRA accuracy. Although IGRAs are primarily indicated for the detection of latent tuberculosis infection, we sought to determine the sensitivity of the TST and 2 whole-blood IGRAs (QuantiFERON-TB assay [QFT] and QuantiFERON-TB Gold assay [QFT-G]) in situations in which infection is confirmed by recovery of M. tuberculosis by culture. METHODS: We conducted a prospective, multicenter, cross-sectional comparison study in which 148 persons suspected to have tuberculosis were tested simultaneously with the TST, QFT, and QFT-G. RESULTS: M. tuberculosis was cultured from samples from 69 (47%) of 148 persons suspected to have tuberculosis; the TST induration was > or = 5 mm for 51 (73.9%) of the 69 subjects (95% confidence interval [CI], 62.5%-82.8%). The QFT indicated tuberculosis infection for 48 (69.6%) of the 69 subjects (95% CI, 57.9%-79.2%) and was indeterminate for 7 (10.1%). The QFT-G yielded positive results for 46 (66.7%) of the 69 subjects (95% CI, 54.9%-76.7%) and indeterminate results for 9 subjects (13.0%). If subjects with indeterminate QFT-G results were excluded, 46 (76.7%) of 60 subjects (95% CI, 64.6%-85.6%) had positive TST results, and the same number of subjects had positive QFT-G results. HIV infection was associated with false-negative TST results but not with false-negative QFT-G results. CONCLUSIONS: The TST, QFT, and QFT-G have similar sensitivity in persons with culture-confirmed infection. As with the TST, negative QFT and QFT-G results should not be used to exclude the diagnosis of tuberculosis in persons with suggestive signs or symptoms.

Simultaneous measurements of cytoplasmic Ca2+ responses and intracellular pH in neutrophils of localized aggressive periodontitis (LAP) patients.

In view of the reports that polymorphonuclear leukocytes (PMN) of patients with localized aggressive periodontitis (LAP) exhibit hyper-responsiveness to stimulation, it has been suggested that such abnormalities could lead to PMN-mediated tissue damage during inflammation. To determine whether these abnormalities include signal transduction, we compared cytoplasmic calcium concentration (Delta[Ca2+](i)) and cytoplasmic pH (DeltapH(i)) changes, early stimulus responses to chemotactic agents, of LAP versus control (C)-PMN and explored whether these could be modulated by sensitizing cytokines or calcium channel-blocking agents. PMN responses of LAP patients were compared with age- and gender-matched controls. Delta[Ca2+](i) and DeltapH(i) were measured fluorimetrically using 1H-indole-6-carboxylic acid, 2-[4-[bis[2-[(acetyloxy)methoxy]-2-oxoethyl]amino]-3-[2-[2-[bis[2-[(acetyloxy)methoxy]-2-oxoethyl]amino]-5-methylphenoxy]ethoxy]phenyl]-1 and 2',7'-bis-(carboxyethyl)-5(6)-carboxyfluorescein as respective probes. Not only was the maximal calcium response to chemoattractants higher in LAP-PMN, but also their subsequent intracellular calcium redistribution was significantly slower. The slower calcium redistribution of LAP-PMN, but not their higher maximal calcium response, was successfully mimicked in C-PMN treated with Nifedipine or 1-[b-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole-HCl, both known to be inhibitors of membrane-associated calcium influx, but this redistribution was not affected when inhibitors of other calcium influx mechanisms, Diltiazem or Verapamil, were used. Taken together, our findings indicate that certain early stimulus responses are aberrant in LAP-PMN, that internal redistribution of cytoplasmic-free calcium is compromised, and, additionally, that a membrane-associated Ca2+ transport defect may be present.

Immune complex stimulation of human neutrophils involves a novel Ca2+/H+ exchanger that participates in the regulation of cytoplasmic pH: flow cytometric analysis of Ca2+/pH responses by subpopulations.

The activation of human phagocytic leukocytes by immune complexes (IC) or opsonized microbes via their Fc and complement receptors has been well-described. The mechanisms involved in this process are complex and depend on the receptors involved. The biochemical events that lead to the destruction of invading organisms in turn display varying degrees of interdependence, but the controlling elements that lead to the ultimate killing of ingested organisms within phagosomes by lysosomal enzymes and reactive oxygen intermediates are still not completely understood. We have addressed these mechanisms by following and correlating the kinetics of responses by individual cells, using multiparameter flow cytometry. Using nonopsonized IC as stimuli, we document here the presence of a novel Ca(2)(+)/H(+) voltage-independent channel in human neutrophils, which helps to control their cytoplasmic pH.