Development and evaluation of specific polymerase chain reaction assays for detecting Theileria equi genotypes.

BACKGROUND: Theileria equi causes equine piroplasmosis, an economically significant disease that affects horses and other equids worldwide. Based on 18S ribosomal RNA (18S rRNA sequences), T. equi can be classified into five genotypes: A, B, C, D, and E. These genotypes have implications for disease management and control. However, no conventional polymerase chain reaction (PCR) assays are available to differentiate the genotypes of T. equi. To overcome this limitation, we developed and evaluated PCR assays specific for the detection of each T. equi genotype. METHODS: A pair of forward and reverse primers, specifically targeting the 18S rRNA sequence of each genotype, was designed. The genotype-specific PCR assays were evaluated for their specificity using plasmids containing inserts of the 18S rRNA sequence of each genotype. Subsequently, the assays were tested on 270 T. equi-positive equine blood DNA samples (92 from donkeys in Sri Lanka and 178 from horses in Paraguay). 18S rRNA sequences derived from the PCR amplicons were analyzed phylogenetically. RESULTS: Each genotype-specific PCR assay accurately targeted the intended genotype, and did not produce any amplicons when 18S rRNA from other T. equi genotypes or genomic DNA of Babesia caballi or uninfected horse blood was used as the template. Previous studies employing PCR sequencing methods identified T. equi genotypes C and D in the Sri Lankan samples, and genotypes A and C in the Paraguayan samples. In contrast, our PCR assay demonstrated exceptional sensitivity by detecting four genotypes (A, C, D, and E) in the Sri Lankan samples and all five genotypes in the Paraguayan samples. All the Sri Lankan samples and 93.3% of the Paraguayan samples tested positive for at least one genotype, further emphasizing the sensitivity of our assays. The PCR assays also had the ability to detect co-infections, where multiple genotypes in various combinations were detected in 90.2% and 22.5% of the Sri Lankan and Paraguayan samples, respectively. Furthermore, the sequences obtained from PCR amplicons clustered in the respective phylogenetic clades for each genotype, validating the specificity of our genotype-specific PCR assays. CONCLUSIONS: The genotype-specific PCR assays developed in the present study are reliable tools for the differential detection of T. equi genotypes.

PCR detection of Theileria equi and Babesia caballi in apparently healthy horses in Paraguay.

Equine piroplasmosis (EP) is a tick-borne disease caused by Theileria equi and Babesia caballi in equids, including horses. EP has a global distribution and often leads to a significant socioeconomic impact on the equine industry. Infected animals remain as carriers and become a source of infection for tick vectors, thereby posing an immense challenge in the disease management. Therefore, detection of these carriers is crucial to assess the risk of transmission and to implement appropriate control measures in endemic countries. Paraguay is a tropical country where various tick-borne diseases are common among livestock; however, the status of EP remains unknown in this country. Because the tick vectors capable of transmitting T. equi and B. caballi are endemic in Paraguay, we hypothesised that Paraguayan horses are infected with these parasite species. To test our hypothesis, we prepared blood DNA samples from a total of 545 apparently healthy horses in 16 of the 17 departments of Paraguay and analysed them with specific PCR assays to detect T. equi and B. caballi. The PCR results showed that 178 (32.7%) and 8 (1.5%) of the horses were infected with T. equi and B. caballi, respectively. Among the infected horses, two (0.4%) were infected with both parasite species. Our analyses further indicated that the positive rates of T. equi infection did not differ between horse breeds, males and females, or age groups. We also found that haematological parameters were the same between the non-infected animals and animals with single infections. By contrast, the two horses co-infected with T. equi and B. caballi had haemoglobin and haematocrit values lower than the normal ranges. In conclusion, the present study demonstrated that Paraguayan horses are infected with T. equi and B. caballi and that the rate of T. equi infection is higher than that of B. caballi. Our findings highlight the need to add EP to the list of differential diagnoses when anaemic horses are presented to equine clinics in Paraguay.

Risk factors for equine trypanosomosis and hematological analysis of horses in Paraguay.

Animal trypanosomosis, caused by Trypanozoon trypanosomes (Trypanosoma evansi and T. equiperdum), and Trypanosoma vivax, is endemic to South American countries and has a negative impact on the livestock industry. However, the risk factors for trypanosomosis in Paraguay remain unknown. This study aimed to determine the risk factors for equine trypanosomosis in Paraguay based on a PCR-based molecular survey and individual horse sampling data. In this study, 739 blood samples were collected from horses in 16 departments of Paraguay between August 2019 and November 2020. To elucidate the risk factors for trypanosome infection, the relationship between trypanosome infection status detected by PCR and the location, sex, age, breed of horses, and season of sample collection was analyzed. There were no significant differences in trypanosome prevalence in horses between the eastern and western regions, ages, or breeds of horses in Paraguay. Sex and season were identified as risk factors for trypanosome infection in horses in Paraguay in the current study. These results suggest that the rainy-summer season, when vectors increase in number and their blood-sucking activity, could be the most important risk factor for trypanosome infection in Paraguay horses. Preventive measures and treatments should be developed to address these factors.

U.S. bound journey of migrant peoples InTransit across Dante's Inferno and Purgatory in the Americas.

Rapid rise of population migration is a defining feature of the 21st century due to the impact of climate change, political instability, and socioeconomic downturn. Over the last decade, an increasing number of migrant peoples travel across the Americas to reach the United States seeking asylum or cross the border undocumented in search of economic opportunities. In this journey, migrant people experience violations of their human rights, hunger, illness, violence and have limited access to medical care. In the 'Divine Comedy', the Italian poet Dante Alighieri depicts his allegorical pilgrimage across Hell and Purgatory to reach Paradise. More than 700 years after its publication, Dante's poem speaks to the present time and the perilious journey of migrant peoples to reach safehavens. By exploring the depths and heights of the human condition, Dante's struggles resonate with the multiple barriers and the unfathomable experiences faced by migrant peoples in transit across South, Central, and North America to reach the United States. Ensuring the safety of migrant peoples across the Americas and elsewhere, and attending to their health needs during their migratory paths represent modern priorities to reduce social injustices and achieving health equity.

First molecular survey of animal trypanosomes in Paraguayan horses.

Despite the epidemic situation of animal trypanosomosis caused by Trypanosoma evansi, Trypanosoma equiperdum and Trypanosoma vivax in South American countries, there are no reports for the prevalence of animal trypanosomes in Paraguay. In this study, 408 blood samples were obtained from apparently healthy horses from sixteen departments of Paraguay, for routine medical check-up from August to September 2019, and a polymerase chain reaction (PCR)-based cross-sectional study was carried out to identify trypanosome prevalence. The prevalence of Trypanozoon (T. evansi and T. equiperdum) and T. vivax was 7.11% (29/408) and 26.23% (107/408), respectively. Mixed infections were detected in 4.90% (20/408) of the samples. Some of the selected trypanosome positive samples were confirmed as T. vivax and T. evansi Type A by sequence analysis of the internal transcribe spacer region and RoTat1.2 variant surface glycoprotein gene, respectively. In conclusion, we found higher prevalence of T. vivax than Trypanozoon in Paraguayan horses. However, the genotypic variation should be verified in further studies.

COVID-19 is associated with higher risk of venous thrombosis, but not arterial thrombosis, compared with influenza: Insights from a large US cohort

IntroductionInfection with SARS-CoV-2 is typically compared with influenza to contextualize its health risks. SARS-CoV-2 has been linked with coagulation disturbances including arterial thrombosis, leading to considerable interest in antithrombotic therapy for Coronavirus Disease 2019 (COVID-19). However, the independent thromboembolic risk of SARS-CoV-2 infection compared with influenza remains incompletely understood. We evaluated the adjusted risks of thromboembolic events after a diagnosis of COVID-19 compared with influenza in a large retrospective cohort. MethodsWe used a US-based electronic health record (EHR) dataset linked with insurance claims to identify adults diagnosed with COVID-19 between April 1, 2020 and October 31, 2020. We identified influenza patients diagnosed between October 1, 2018 and April 31, 2019. Primary outcomes [venous composite of pulmonary embolism (PE) and acute deep vein thrombosis (DVT); arterial composite of ischemic stroke and myocardial infarction (MI)] and secondary outcomes were assessed 90 days post-diagnosis. Propensity scores (PS) were calculated using demographic, clinical, and medication variables. PS-adjusted hazard ratios (HRs) were calculated using Cox proportional hazards regression. ResultsThere were 417,975 COVID-19 patients (median age 57y, 61% women), and 345,934 influenza patients (median age 47y, 66% women). Compared with influenza, patients with COVID-19 had higher venous thromboembolic risk (HR 1.53, 95% CI 1.38-1.70), but not arterial thromboembolic risk (HR 1.02, 95% CI 0.95-1.10). Secondary analyses demonstrated similar risk for ischemic stroke (HR 1.11, 95% CI 0.98-1.25) and MI (HR 0.93, 95% CI 0.85-1.03) and higher risk for DVT (HR 1.36, 95% CI 1.19-1.56) and PE (HR 1.82, 95% CI 1.57-2.10) in patients with COVID-19. ConclusionIn a large retrospective US cohort, COVID-19 was independently associated with higher 90-day risk for venous thrombosis, but not arterial thrombosis, as compared with influenza. These findings may inform crucial knowledge gaps regarding the specific thromboembolic risks of COVID-19.

Guía de Práctica Clínica de enfermedad herniaria inguinal de la niñez / Clinical Practice Guidelines children´s inguinal hernia disease

La enfermedad herniaria inguinal de la niñez, comprende un grupo de afecciones de la región inguinal que tienen su génesis en fallos de la obliteración del conducto peritoneo vaginal o conducto vaginal. El propósito de los autores es compartir esta guía con los cirujanos pediátricos a través de su publicación, lo que les permitiría emplearla como referencia en sus instituciones. La enfermedad herniaria inguinal de la niñez, constituye la primera causa de operaciones electivas ‒programadas o planificadas‒ en unidades quirúrgicas pediátricas. Tiene una incidencia que varía entre 1 y 7 por ciento de la población infantil. En el IV Simposio Nacional de Cirugía Pediátrica (Varadero, Matanzas, 1- 3 de juliode 2019) se presentó, discutió y se aprobó esta guía de práctica clínica. Su aplicación en diferentes servicios de cirugía pediátrica beneficiaría a un gran número de niños con esta enfermedad, además de ser útil como orientación a profesionales encargados de la atención sanitaria a niños y adolescentes en la atención primaria de salud(AU)

Children´s inguinal hernia disease comprises a group of conditions in the groin region that have their genesis in failures of peritoneal vaginal duct obliteration or vaginal duct. The purpose of the authors is to share these guidelines with pediatric surgeons through its publication, which would allow them to use it as a reference in their institutions. Children´s inguinal hernia disease is the leading cause of elective operations, programmed or planned, in pediatric surgical units. It has an incidence ranging from 1 to 7 percent of the children population. At the IV National Symposium on Pediatric Surgery (Varadero, Matanzas, 1-3 July 2019) these clinical practice guidelines were presented, discussed and approved. Their application in different pediatric surgery services would benefit a large number of children with this disease, as well as being useful as guidance to professionals in charge of health care for children and adolescents in the primary health care level(AU)

Association of HIV with COVID-19 Outcomes Among Hospitalized Adults

BackgroundWhether HIV infection is associated with differences in clinical outcomes among people hospitalized with COVID-19 is uncertain. ObjectiveTo evaluate the impact of HIV infection on COVID-19 outcomes among hospitalized patients. MethodsUsing the American Heart Associations COVID-19 Cardiovascular Disease registry, we used hierarchical mixed effects models to assess the association of HIV with in-hospital mortality accounting for patient demographics and comorbidities and clustering by hospital. Secondary outcomes included major adverse cardiac events (MACE), severity of illness, and length of stay (LOS). ResultsThe registry included 21,528 hospitalization records of people with confirmed COVID-19 from 107 hospitals in 2020, including 220 people living with HIV (PLWH). PLWH were younger (56.0+/-13.0 versus 61.3+/-17.9 years old) and more likely to be male (72.3% vs 52.7%), Non-Hispanic Black (51.4% vs 25.4%), on Medicaid (44.5% vs 24.5), and active tobacco users (12.7% versus 6.5%). Of the study population, 36 PLWH (16.4%) had in-hospital mortality compared with 3,290 (15.4%) without HIV (Risk ratio 1.06, 95%CI 0.79-1.43; risk difference 0.9%, 95%CI -4.2 to 6.1%; p=0.71). After adjustment for age, sex, race, and insurance, HIV was not associated with in-hospital mortality (aOR 1.13; 95%CI 0.77-1.6; p 0.54) even after adding body mass index and comorbidities (aOR 1.15; 95%CI 0.78-1.70; p=0.48). HIV was not associated with MACE (aOR 0.99, 95%CI 0.69-1.44, p=0.91), severity of illness (aOR 0.96, 95%CI 0.62-1.50, p=0.86), or LOS (aOR 1.03; 95% CI 0.76-1.66, p=0.21). ConclusionHIV was not associated with adverse outcomes of COVID-19 including in-hospital mortality, MACE, or severity of illness. Condensed AbstractWe studied 21,528 patients hospitalized with COVID-19 at 107 hospitals in AHAs COVID-19 registry to examine the association between HIV and COVID-19 outcomes. More patients with HIV were younger, male, non-Hispanic Black, on Medicaid and current smokers. HIV was not associated with worse COVID-19 in-hospital mortality (Risk ratio 1.06, 95%CI 0.79-1.43; p=0.71) even after adjustment (aOR 1.15; 95%CI 0.78-1.70; p=0.48). HIV was also not associated with MACE (aOR 0.99, 95%CI 0.69-1.44, p=0.91) or severity of illness (aOR 0.96, 95%CI 0.62-1.50, p=0.86. Our findings do not support that HIV is a major risk factor for adverse COVID-19 outcomes.

Galectin-3 as a potential prognostic biomarker of severe COVID-19 in SARS-CoV-2 infected patients

BACKGROUNDPrognostic biomarkers are needed to identify patients at high-risk for severe COVID-19. Galectin-3 is known to drive neutrophil infiltration and release of pro-inflammatory cytokines contributing to airway inflammation. METHODSIn this prospective cohort, we assessed galectin-3 levels in 156 hospitalized patients with confirmed COVID-19. COVID-19 patients were diagnosed as either critical (>50% lung damage) or moderate (<50% of lung damage) based on computerized tomography. Patients who required invasive mechanical ventilation (IMV) and/or died during hospitalization were categorized as having a severe outcome, and a non-severe outcome if they were discharged and none of the former occurred. RESULTSElevated serum galectin-3 was significantly higher in critical patients compared to moderate ones (35.91 {+/-} 19.37 ng/mL vs. 25 {+/-} 14.85 ng/mL, p<0.0001). Patients who progressed to a severe outcome including IMV and/or in-hospital death, presented higher galectin-3 levels (41.17 ng/mL [IQR 29.71 - 52.25] vs. 23.76 ng/mL [IQR 15.78 - 33.97] compared to those of a non-severe outcome, p<0.0001). Galectin-3 discriminated well between those with severe and non-severe outcome, with an AUC of 0.75 (95% CI 0.67 - 0.84, p<0.0001) and was found to be an independent predictor of severe outcome regardless of the percentage of lung involvement. Additionally, the combination of galectin-3, CRP and albumin, significantly improved its individual predicting ability with an AUC 0.84 (95% CI 0.77 - 0.91, p<0.0001). CONCLUSIONCirculating galectin-3 levels can be used to predict severe outcomes in COVID-19 patients, including the requirement of mechanical ventilation and/or death, regardless of the initial severity of the disease.

Improving the antiviral efficacy and selectivity of HIV-1 reverse transcriptase inhibitor TSAO-T by the introduction of functional groups at the N-3 position.

Novel derivatives of the anti-HIV-1 agent, TSAO-T, bearing at the N-3 position a variety of polar, lipophilic, or aromatic groups linked to that position through flexible polymethylene linkers of different length, were prepared and evaluated for their anti-HIV activity. Several compounds (within the series of polar bearing groups) exhibited a 2-6-fold improved antiviral potency with regard to the lead compound, TSAO-T. Moreover, some of the most active N-3 TSAO derivatives retain antiviral activity against the TSAO-T-resistant HIV-1 strain (Glu138 --> Lys). Interestingly, the N-methylcarboxamide derivative 17 was 5- to 6-fold more active (IC50: 0.56 microM) against recombinant HIV-1 reverse transcriptase than the lead compound, thus becoming the most active TSAO derivative synthesized so far. On the other hand, the N-3 methylcarboxamide TSAO derivative 12 turned out to have the highest selectivity index yet reported for this class of compounds (around > or =12 000).