Blood Polymerase Chain Reaction Research Articles

Zoonosis screening in Spanish immunocompromised children and their pets.

Although pets provide several social-emotional benefits for children, the risk of zoonosis must be considered among immunocompromised individuals. A prospective study was conducted in a tertiary hospital including immunocompromised patients younger than 20 years owning dogs and/or cats. Colonization and/or infection was evaluated by stool studies, bacterial swabs, blood polymerase chain reaction and serological studies in both patients and their pets, to evaluate potential zoonotic transmission occurrence. We included 74 patients and their 92 pets (63 dogs, 29 cats). Up to 44.6% of the patients and 31.5% of the pets had at least 1 positive result. Up to 18.4% of pets' fecal samples were positive (bacteria, parasites or hepatitis E virus). No helminths were observed despite the high frequency of incorrect intestinal deworming practices. Among children, gastrointestinal microorganisms were found in 37.3% (primarily Clostridium difficile). Colonization by Staphylococcus pseudintermedius was common among pets (8.0%) but not among children (0.0%). No shared colonization between owners and pets was observed, except in one case (Blastocystis in both patient and pet feces). Among patients, serologies were positive for Strongyloides stercoralis (14.8%), Toxocara canis (3.2%), Bartonella henselae (19.1%) and hepatitis E (5.6%). Serology was positive for Rickettsia spp. (22.6%) and Babesia spp. (6.5%) in dogs and for Leishmania spp. (14.3%) and Toxoplasma spp. (14.3%) in cats. Exposure to zoonotic agents was detected in both patients and pets; however, shared colonization events were almost nonexistent. In our cohort, dogs and cats do not appear to entail high zoonosis transmission risk for immunocompromised patients.

Successful treatment of cicatricial stenosis of the main, intermediate and upper lobe bronchi of the right lung complicated by total lung atelectasis in an infant: a clinical case

BACKGROUND: The authors present a rare clinical case of stenosis of the main, intermediate and upper lobe bronchi of the right lung complicated by total lung atelectasis in an infant. They discuss issues of complex disease diagnostics, analyze X-ray and bronchological images, assess outcomes after multidisciplinary approach in the treatment of a premature baby as well as analyze data of catamnestic observation. CLINICAL CASE DESCRIPTION: Girl T., one of monochorionic diamniotic twins, was born at 27 week gestation in 2021 with weight at birth 825 grams, Apgar score 4/5. Since her birth, she had artificial lung ventilation due to her deep immaturity and developed severe respiratory distress syndrome. At the age of 19 days, total atelectasis of the right lung was diagnosed in the child as a result of stenosis formation in the main, intermediate and upper lobe bronchi which was confirmed by clinical and radiological findings. A probable etiology of acquired stenosis is consequences of prolonged intubation under insufficiently formed cartilaginous framework. Chlamydial infection, detected in the child, could also support inflammatory process in the bronchi. The child was examined with the following instruments: radiography, multispiral computed tomography-angiography of the thoracic cavity, bronchofibroscopy, echocardiography. Laboratory tests: polymerase chain reaction of blood and sputum for herpes viruses type 1–6, mycoplasma, ureaplasma, Chlamydia pneumonia; blood enzyme immunoassays — antibodies to viruses of Herpesviridae type 1–6, Chlamydia trachomatis. Conservative therapy provided only temporal improvement; atelectasis was relapsing. At the age of 5 months and 20 days, the child underwent bronchoplasty with resection of damaged areas of the bronchi. Further, age-related diameter of the bronchial lumen was maintained by the staged bougienage. Catamnestic observation lasted for 3 years. Late timing of surgical intervention is explained by severe problems in a deeply premature and immature child that required immediate treatment (intraventricular hemorrhages, hydrocephalus, retinopathy, etc.). Effective surgical treatment (bronchoplasty), active growth of lung tissue at the first year of life as well as reasonable rehabilitation allowed to obtain good long-term outcomes. By the age of 1 year 2 months, normalization of right lung airiness was achieved. Currently, there is a slight reduction in right lung volume. Multispiral computed tomography-angiography revealed a few cylindrical bronchoectases in the segmental and subsegmental bronchi of the upper and middle lobes of the right lung which do not have clinical equivalents yet. The child is under pulmonologist's observation and management. CONCLUSION: A multidisciplinary approach to the treatment of complicated patient's state and high compensatory capabilities of neonatal and infantile periods have promoted the restoration of normal lung aeration and functioning after long-lasting lung atelectasis

Effects of short-chain fatty acid-butyrate supplementation on expression of circadian-clock genes, sleep quality, and inflammation in patients with active ulcerative colitis: a double-blind randomized controlled trial

BackgroundThe regulation of the circadian clock genes, which coordinate the activity of the immune system, is disturbed in inflammatory bowel disease (IBD). Emerging evidence suggests that butyrate, a short-chain fatty acid produced by the gut microbiota is involved in the regulation of inflammatory responses as well as circadian-clock genes. This study was conducted to investigate the effects of sodium-butyrate supplementation on the expression of circadian-clock genes, inflammation, sleep and life quality in active ulcerative colitis (UC) patients.MethodsIn the current randomized placebo-controlled trial, 36 active UC patients were randomly divided to receive sodium-butyrate (600 mg/kg) or placebo for 12-weeks. In this study the expression of circadian clock genes (CRY1, CRY2, PER1, PER2, BMAl1 and CLOCK) were assessed by real time polymerase chain reaction (qPCR) in whole blood. Gene expression changes were presented as fold changes in expression (2^-ΔΔCT) relative to the baseline. The faecal calprotectin and serum level of high-sensitivity C-reactive protein (hs-CRP) were assessed by enzyme-linked immunosorbent assay method (ELIZA). Moreover, the sleep quality and IBD quality of life (QoL) were assessed by Pittsburgh sleep quality index (PSQI) and inflammatory bowel disease questionnaire-9 (IBDQ-9) respectively before and after the intervention.ResultsThe results showed that sodium-butyrate supplementation in comparison with placebo significantly decreased the level of calprotectin (-133.82 ± 155.62 vs. 51.58 ± 95.57, P-value < 0.001) and hs-CRP (-0.36 (-1.57, -0.05) vs. 0.48 (-0.09-4.77), P-value < 0.001) and upregulated the fold change expression of CRY1 (2.22 ± 1.59 vs. 0.63 ± 0.49, P-value < 0.001), CRY2 (2.15 ± 1.26 vs. 0.93 ± 0.80, P-value = 0.001), PER1 (1.86 ± 1.77 vs. 0.65 ± 0.48, P-value = 0.005), BMAL1 (1.85 ± 0.97 vs. 0.86 ± 0.63, P-value = 0.003). Also, sodium-butyrate caused an improvement in the sleep quality (PSQI score: -2.94 ± 3.50 vs. 1.16 ± 3.61, P-value < 0.001) and QoL (IBDQ-9: 17.00 ± 11.36 vs. -3.50 ± 6.87, P-value < 0.001).ConclusionButyrate may be an effective adjunct treatment for active UC patients by reducing biomarkers of inflammation, upregulation of circadian-clock genes and improving sleep quality and QoL.

A commercial whole blood polymerase chain reaction assay failed to diagnose histoplasmosis in cats confirmed by cytology or urine antigen detection.

To determine the sensitivity and specificity of a commercial whole blood real-time PCR assay (RT-PCR) for the diagnosis of histoplasmosis when compared to direct organism identification and/or urine antigen quantification by enzyme immunoassay (UA-EIA). A secondary objective was to compare the sensitivity and specificity of RT-PCR to anti-Histoplasma immunoglobulin G antibody detection by enzyme immunoassay (IgG-EIA) and IgG-EIA to UA-EIA. Cats presented to the Kansas State University Veterinary Health Center from February through September of 2023 in which histoplasmosis was diagnosed or suspected. From February through September of 2023, cats were tested by RT-PCR, IgG-EIA, and UA-EIA if histoplasmosis was diagnosed cytologically or was a differential diagnosis for the presenting clinical signs. Cats were excluded if all 3 tests were not submitted or if the diagnosis of histoplasmosis could not be excluded despite a negative UA-EIA result. Cats with cytologically or histologically confirmed histoplasmosis were designated as proven histoplasmosis cases, and cats with a positive UA-EIA result without cytological or histological confirmation were designated as probable histoplasmosis cases. 10 cats were diagnosed with either proven (n = 6) or probable (4) histoplasmosis, and 10 cats were considered true negatives. Whole blood RT-PCR results were negative in all 20 cats (sensitivity, 0%; 95% CI, 0% to 30.85%). The IgG-EIA was 90% sensitive (95% CI, 55.50% to 99.75%) and 70% specific (95% CI, 34.75% to 93.33%). The UA-EIA results were positive in all cats with proven histoplasmosis. This commercial RT-PCR is insensitive when used on whole blood collected in EDTA and should not be used to diagnose feline histoplasmosis. Further studies are required to determine whether alternate RT-PCR protocols for EDTA-collected whole blood could be useful for diagnosing histoplasmosis in cats.

Etiology of Acute Febrile Illnesses in Adults in the Defense Community in French Guiana.

In tropical countries, acute febrile illnesses represent a complex clinical problem for general practitioners. We describe the prevalence of different etiologies of acute febrile illnesses occurring among French service members and their families, excluding children, in general practice in French Guiana. From June 2017 to March 2020, patients with a fever ≥ 37.8°C with a duration of less than 15 days who sought medical care at the army medical centers in Cayenne and Kourou were prospectively enrolled. Based on clinical presentation, blood, urine, nasopharyngeal, and stool samples were collected for diagnostic testing for viruses, bacteria, and parasites (by direct examination, microscopic examination of blood smears, culture, serology, or polymerase chain reaction), and standardized biological tests were systematically performed. Among 175 patients retained for analysis, fever with nonspecific symptoms was predominant (46.9%), with 10 Plasmodium vivax malaria cases, 8 dengue infections, and 6 cases of Q fever. The second most frequent cause of acute febrile illness was upper respiratory tract infections (32.0%) due to influenza virus (n = 18) or human rhinovirus (n = 10). Among the causes of acute febrile illness in French Guiana, clinicians should first consider arboviruses and malaria, as well as Q fever in cases of elevated C-reactive protein with nonspecific symptoms and influenza in cases of signs and symptoms associated with upper respiratory tract infections. Despite an expanded microbiological search, the etiology of 51.4% of acute febrile illnesses remain unknown. Further investigations will be necessary to identify the etiology of acute febrile illnesses, including new pathogens, in French Guiana.

CLOCK gene circannual expression in cluster headache.

Cluster headache is a primary headache disorder characterized by bouts with circadian and circannual patterns. The CLOCK gene has a central role in regulating circadian rhythms. Here, we investigate the circannual CLOCK expression in a population of cluster headache patients in comparison to matched controls. Patients with cluster headache were sampled two to four times over at least one year, both in or outside bouts, one week after each solstice and equinox. The expression of CLOCK was measured by quantitative real-time polymerase chain reaction (RT-PCR) in the peripheral blood. This study included 50 patients and 58 matched controls. Among the patient population, composed of 42/50 males (84%) with an average age of 44.6 years, 45/50 (90%) suffered from episodic cluster headache. Two to four samples were collected from each patient adding up to 161 samples, 36 (22.3%) of which were collected within a bout. CLOCK expression for cluster headache patients was considerably different from that of the control population in winter (p-value mean = 0.006283), spring (p-value mean = 0.000006) and summer (p-value mean = 0.000064), but not in autumn (p-value mean = 0.262272). For each season transition, the variations in CLOCK expression were more pronounced in the control group than in the cluster headache population. No statistically significant differences were found between bout and non-bout samples. No individual factors (age, sex, circadian chronotype, smoking and coffee habits or history of migraine) were related to CLOCK expression. We observed that CLOCK expression in cluster headache patients fluctuates less throughout the year than in the control population. Bout activity and lifestyle factors do not seem to influence CLOCK expression.

Investigation of Trypanosoma evansi in Sumatran Elephant (Elephas maximus sumatranus) in Indonesia Using Various Methods

Trypanosoma evansi is a widespread hemoprotozoa that causes trypanosomiasis in both livestock and wildlife. One of the susceptible animals is the Sumatran elephant, which is included in the endangered species category. The infection of this parasite often develops into chronic and sub-clinical forms in elephants, therefore it may become unnoticed and hard to diagnose. This study aimed to analyze and evaluate the infection of T. evansi in semi-captive Sumatran elephants in Way Kambas National Park, Indonesia, using various diagnostic tests. The prevalence of T. evansi in a total of 53 Sumatran elephants was estimated using a card agglutination test for trypanosomiasis (CATT) in 2016. A longitudinal study was later conducted in 2019 using Giemsa stained blood smear (GSBS) and polymerase chain reaction (PCR). The results showed that 26.4% of samples (14/53) were positively detected by both CATT in 2016 and PCR in 2019, while GSBS was unable to detect the parasites in all samples. Furthermore, four individuals were confirmed to have persistent infections. This study concluded that the ability of CATT and PCR were more convincing over GSBS for the diagnosis of sub-clinical trypanosomiasis in Sumatran elephants. However, it is recommended to use a combination of CATT as a screening tool and PCR as a confirmatory test for reliable results.

Diagnosis of human leptospirosis: systematic review and meta-analysis of the diagnostic accuracy of the Leptospira microscopic agglutination test, PCR targeting Lfb1, and IgM ELISA to Leptospira fainei serovar Hurstbridge

BackgroundLeptospirosis is an underdiagnosed infectious disease with non-specific clinical presentation that requires laboratory confirmation for diagnosis. The serologic reference standard remains the microscopic agglutination test (MAT) on paired serum samples. However, reported estimates of MAT’s sensitivity vary. We evaluated the accuracy of four index tests, MAT on paired samples as well as alternative standards for leptospirosis diagnosis: MAT on single acute-phase samples, polymerase chain reaction (PCR) with the target gene Lfb1, and ELISA IgM with Leptospira fainei serovar Hurstbridge as an antigen.MethodsWe performed a systematic review of studies reporting results of leptospirosis diagnostic tests. We searched eight electronic databases and selected studies that tested human blood samples and compared index tests with blood culture and/or PCR and/or MAT (comparator tests). For MAT selection criteria we defined a threshold for single acute-phase samples according to a national classification of leptospirosis endemicity. We used a Bayesian random-effect meta-analysis to estimate the sensitivity and specificity of MAT in single acute-phase and paired samples separately, and assessed risk of bias using the Quality Assessment of Studies of Diagnostic Accuracy Approach- 2 (QUADAS-2) tool.ResultsFor the MAT accuracy evaluation, 15 studies were included, 11 with single acute-phase serum, and 12 with paired sera. Two included studies used PCR targeting the Lfb1 gene, and one included study used IgM ELISA with Leptospira fainei serovar Hurstbridge as antigen. For MAT in single acute-phase samples, the pooled sensitivity and specificity were 14% (95% credible interval [CrI] 3–38%) and 86% (95% CrI 59–96%), respectively, and the predicted sensitivity and specificity were 14% (95% CrI 0–90%) and 86% (95% CrI 9–100%). Among paired MAT samples, the pooled sensitivity and specificity were 68% (95% CrI 32–92%) and 75% (95% CrI 45–93%) respectively, and the predicted sensitivity and specificity were 69% (95% CrI 2–100%) and 75% (2–100%).ConclusionsBased on our analysis, the accuracy of MAT in paired samples was not high, but it remains the reference standard until a more accurate diagnostic test is developed. Future studies that include larger numbers of participants with paired samples will improve the certainty of accuracy estimates.

Epidemiology and treatment of invasive Bartonella spp. infections in the United States.

Bartonella spp., renowned for cat-scratch disease, has limited reports of dissemination. Tissue and blood cultures have limitations in detecting this fastidious pathogen. Molecular testing (polymerase chain reaction, PCR) and cell-free DNA have provided an avenue for diagnoses.This retrospective observational multicenter study describes the incidence of disseminated Bartonella spp. and treatment-related outcomes. Inclusion criteria were diagnosis of bartonellosis via diagnosis code, serology testing of blood, polymerase chain reaction (PCR) of blood, 16/18S tests of blood or tissue, cultures of blood or tissue, or cell-free DNA of blood or tissue from January 1, 2014, through September 1, 2021. Exclusions were patients who did not receive treatment, insufficient data on treatment course, absence of dissemination, or retinitis as dissemination. Patients were primarily male (n = 25, 61.0%), white (n = 28, 68.3%), with mean age of 50 years (SD 14.4), and mean Charlson comorbidity index of 3.5 (SD 2.1). Diagnosis was primarily by serology (n = 34, 82.9%), with Bartonella henselae (n = 40, 97.6%) as the causative pathogen. Treatment was principally doxycycline with rifampin (n = 17, 41.5%). Treatment failure occurred in 16 (39.0%) patients, due to escalation of therapy during treatment (n = 5, 31.3%) or discontinuation of therapy due to an adverse event or tolerability (n = 5, 31.3%). In conclusion, this is the largest United States-based cohort of disseminated Bartonella spp. infections to date with a reported 39% treatment failure. This adds to literature supporting obtaining multiple diagnostic tests when Bartonella is suspected and describes treatment options.

Chronic High-Level Parasitemia in HIV-Infected Individuals With or Without Visceral Leishmaniasis in an Endemic Area in Northwest Ethiopia: Potential Superspreaders?

People with human immunodeficiency virus (PWH) with recurrent visceral leishmaniasis (VL) could potentially drive Leishmania transmission in areas with anthroponotic transmission such as East Africa, but studies are lacking. Leishmania parasitemia has been used as proxy for infectiousness. This study is nested within the Predicting Visceral Leishmaniasis in HIV-InfectedPatients (PreLeisH) prospective cohort study, following 490 PWH free of VL at enrollment for up to 24-37 months in northwest Ethiopia. Blood Leishmania polymerase chain reaction (PCR) was done systematically. This case series reports on 10 PWH with chronic VL (≥3 VL episodes during follow-up) for up to 37 months, and 3 individuals with asymptomatic Leishmania infection for up to 24 months. All 10 chronic VL cases were male, on antiretroviral treatment, with 0-11 relapses before enrollment. Median baseline CD4 count was 82 cells/µL. They displayed 3-6 VL treatment episodes over a period up to 37 months. Leishmania blood PCR levels were strongly positive for almost the entire follow-up (median cycle threshold value, 26 [interquartile range, 23-30]), including during periods between VL treatment. Additionally, we describe 3 PWH with asymptomatic Leishmania infection and without VL history, with equally strong Leishmania parasitemia over a period of up to 24 months without developing VL. All were on antiretroviral treatment at enrollment, with baseline CD4 counts ranging from 78 to 350 cells/µL. These are the first data on chronic parasitemia in PWH from Leishmania donovani-endemic areas. PWH with asymptomatic and symptomatic Leishmania infection could potentially be highly infectious and constitute Leishmania superspreaders. Xenodiagnosis studies are required to confirm infectiousness.

Disparities in Cytomegalovirus Infection Rates by Race and Ethnicity among Pediatric Allogeneic Hematopoietic Cell Transplantation Recipients at a Single Center

Previous literature has reported cytomegalovirus (CMV) infection rate disparities among racial/ethnic groups of hematopoietic cell transplantation (HCT) recipients. Because race and ethnicity categorizations are social constructs unlikely to affect biological systems, it is likely there are covariates on the pathway to CMV detection, known as mediators, that can explain the observed disparity. Recent developments in mediation analysis methods enable the analysis of time-to-event outcomes, allowing an investigation of these disparities to also consider the timing of CMV infection detection relative to HCT. This study aimed to explore whether racial and ethnic CMV infection disparities existed within a population of HCT recipients at our center, and whether clinical covariates explained any observed association. The study cohort included all recipients of allogeneic HCT performed at the Children's Hospital of Philadelphia between January 2004 and April 2017 who were CMV PCR-negative pretransplantation, had known donor/recipient CMV serology, and were under blood CMV PCR surveillance. Subjects were followed for 100 days post-HCT. Accelerated failure time models using subject's reported race/ethnicity, dichotomized into non-Hispanic White (NHW) and non-NHW, and exposure and time to CMV detection as outcomes examined whether selected clinical factors-donor/recipient CMV serostatus, recipient age, indication for HCT, hematopoietic cell source, match quality-mediated any identified exposure-outcome association. The analysis included 348 HCTs performed in 335 subjects, with 86 episodes (24.7%) in which CMV was detected via PCR analysis. The accelerated failure time model without mediators estimated that non-NHW subjects had fewer CMV-free survival days (time ratio, .21; 95% confidence interval, .10 to .44). Any hypothesized mediator mediated at most 5% of the total association between race/ethnicity and time to CMV detection. Non-NHW HCT recipients had fewer CMV-free survival days than NHW recipients; none of the clinical factors hypothesized to mediate this association accounted for a significant component of total association. Further research should focus on nonclinical factors influenced by systemic racism to better understand their effect on CMV infection among HCT recipients.

An algorithm for visceral leishmaniasis diagnosis in Brazil from a 10-year analysis of National Reference Laboratory data.

Visceral leishmaniasis (VL) requires diagnostic assays to complement clinical suspicion. However, there is no standardization of a diagnostic flow using available assays. This study aimed to evaluate the performance of parasitological, molecular, and serological assays for diagnosing VL and propose a diagnostic flow based on performance, practicality, and invasiveness. We conducted a study of 10-year (2010-2020) routine diagnoses of VL at the Brazilian National Reference Laboratory. We propose a diagnostic flow where individuals suspected of VL are initially screened for antibodies using an immunochromatographic test (ICT) with rK39 antigen on the nitrocellulose membrane. This is followed by a blood polymerase chain reaction (PCR) for Leishmania sp. kDNA and direct parasitological exam and/or PCR in bone marrow aspirate. A positive result in any of these assays can define a VL case. If clinical suspicion persists in negative individuals, the diagnostic flow should be repeated. The proposed flow has the potential to standardize and improve the diagnosis of VL. It reduces the need for invasive tests without compromising diagnostic accuracy.

Clinical Characteristics of Infants with Symptomatic Congenital and Postnatal Cytomegalovirus Infection-An 11-Year Multicenter Cohort Study in Taiwan.

(1) Background: Cytomegalovirus (CMV) infection is a prevalent viral disease among infants. The prevalence typically ranges from 0.2% to 2.4% among all newborns. There are limited data regarding the demographic characteristics of infants with symptomatic CMV infections. (2) Methods: In this retrospective cohort study using the Chang Gung Memorial Hospital multicenter database, infants with CMV infection determined by a positive urine culture, positive blood polymerase chain reaction assay or positive immunoglobulin M result for CMV from 2011 through 2021 were included. Clinical characteristics at initial diagnosis, management and outcomes were investigated. Congenital CMV (cCMV) infection is diagnosed within three weeks after birth; postnatal CMV (pCMV) is diagnosed when CMV is detected after the first 3 weeks of life. (3) Results: Among the 505 CMV-infected infants identified, 272 were included in the analysis. According to the age at initial presentation, 21 infants had cCMV infection and 251 had pCMV infection. Higher incidences of prematurity and being small for gestational age and a lower Z score for weight at diagnosis were observed in the cCMV group. While thrombocytopenia (61.9%) was the leading presentation in the cCMV group, hepatitis (59.8%) and prolonged jaundice (21.9%) were more common in the pCMV group. (4) Conclusions: Utilizing an 11-year multicenter database, we demonstrated the characteristics of infants with CMV infection in Taiwan and highlighted the demographic disparities and differing symptoms between the cCMV and pCMV groups. These findings emphasize the necessity for future research to refine screening policies, explore treatment options, and establish follow-up protocols for affected infants.

Five years of post-validation surveillance of lymphatic filariasis in Thailand

BackgroundThe World Health Organization (WHO) validated Thailand in 2017 as having eliminated lymphatic filariasis (LF) as a public health problem with recommendations for continued surveillance. This article describes measures and progress made in Thailand with post-validation surveillance (PVS) of LF from 2018 until 2022.MethodsThe implementation unit (IU) is a sub-village in 11 former LF endemic provinces. Human blood surveys are targeted in 10% of IUs each year. In Wuchereria bancrofti areas, filaria antigen test strips (FTS) are used, and in Brugia malayi areas, antibody test kits (Filaria DIAG RAPID) are used. Positive cases are confirmed by thick blood film (TBF) and polymerase chain reaction (PCR). Vector surveys for mosquito species identification and dissection for microfilaria (Mf)/filarial larvae are done in 1% of IUs where human blood surveys are conducted. Human blood surveys using FTS are conducted among migrants in five provinces. Surveillance of cats is done in areas that previously recorded > 1.0% Mf rate among cats. Morbidity management and disability prevention (MMDP) are done every 2 years in LF-endemic areas where chronic disease patients reside.ResultsFrom 2018 to 2022, in a total of 357 IUs in 11 provinces, human blood surveys were conducted in 145 IUs (41%) with an average population coverage of 81%. A total of 22,468 FTS and 27,741 FilariaDIAG RAPID were performed. 27 cases were detected: 3 cases of W. bancrofti in Kanchanaburi province and 24 cases of B. malayi in Narathiwat province. 4 cases of W. bancrofti were detected in two provinces through routine public health surveillance. Vector surveys in 47 IUs detected B. malayi Mf filarial larvae only in Narathiwat province. Chronic LF patients reduced from 114 in 2017 to 76 in 2022. Surveys among 7633 unregistered migrants yielded 12 cases of W. bancrofti. Mf rate among cats in Narathiwat province declined from 1.9% in 2018 to 0.7% in 2022. MMDP assessments revealed gaps in healthcare provider’s management of chronic cases due to staff turnover.ConclusionsIn 2022, after 5 years of PVS, Thailand re-surveyed 41% of its previously endemic IUs and demonstrated ongoing transmission in only one province of Narathiwat, where Mf prevalence is below the WHO provisional transmission threshold of 1%. This study highlights the importance of continued disease surveillance measures and vigilance among health care providers in LF receptive areas.Graphical

Estimation of parasitaemia in imported falciparum malaria using the results of a combined rapid diagnostic test. No big help from haematological parameters

BackgroundMicroscopy continues to be the mainstay for the evaluation of parasitaemia in malaria but requires laboratory support and microbiological experience. Other fast and simple methods are necessary.MethodsA retrospective observational study of imported malaria treated from July-2007 to December-2020 was carried out to evaluate the association between the degree of parasitaemia and both rapid diagnostic tests (RDT) reactivity patterns and haematological parameters. Plasmodium falciparum monoinfections diagnosed by peripheral blood smear and/or polymerase chain reaction (PCR),which also had a positive RDT result in the same blood sample, were included in the study.ResultsA total of 273 patients were included. Most of them were male (n = 256; 93.8%) and visiting friends and relatives (VFR) travellers (n = 252; 92.3%). Patients with plasmodial lactate dehydrogenase (pLDH) or aldolase and histidine-rich protein 2 (HRP-2) co-reactivity (Pan/Pf pattern) had a parasitaemia range between 0 and 37% while those with just HRP-2 reactivity (P. falciparum pattern) had ranges between 0 and 1%. Not a single case of P. falciparum pattern was found for parasitaemia ranges greater than 1%, showing a negative predictive value of 100% for high parasitaemia. All the correlations between haematological parameters and parasitaemia resulted to be weak, with a maximum rho coefficient of -0.35 for lymphocytes and platelets, and of 0.40 for neutrophils-to-lymphocytes count ratio. Multivariate predictive models were constructed reflecting a poor predictive capacity.ConclusionsThe reactivity pattern of RDT allows a rapid semi-quantitative assessment of P. falciparum parasitaemia in travellers with imported malaria, discriminating patients with lower parasite loads. Haematological parameters were not able to estimate parasitaemia with sufficient precision.

A new genotype of hepatitis A virus causing transient liver enzyme elevations in Mauritius-origin laboratory-housed Macaca fascicularis.

Hepatitis A virus (HAV) infects humans and nonhuman primates, typically causing an acute self-limited illness. Three HAV genotypes have been described so far for humans, and three genotypes have been described for nonhuman primates. We observed transiently elevated liver enzymes in Mauritius-origin laboratory-housed macaques in Germany and were not able to demonstrate an etiology including HAV by serology and polymerase chain reaction (PCR). HAV is a rare pathogen in cynomolgus macaques, and since all employees were routinely vaccinated against HAV, it was not a part of the routine vaccination and screening program. A deep sequencing approach identified a new HAV genotype (referred to as Simian_HAV_Macaca/Germany/Mue-1/2022) in blood samples from affected animals. This HAV was demonstrated by reverse transcription PCR in blood and liver and by in situ hybridization in liver, gall bladder, and septal ducts. A commercial vaccine was used to protect animals from liver enzyme elevation. The newly identified simian HAV genotype demonstrates 80% nucleotide sequence identity to other simian and human HAV genotypes. There was deeper divergence between Simian_HAV_Macaca/Germany/Mue-1/2022 and other previously described HAVs, including both human and simian viruses. In situ hybridization indicated persistence in the biliary epithelium up to 3 months after liver enzymes were elevated. Vaccination using a commercial vaccine against human HAV prevented reoccurrence of liver enzyme elevations. Because available assays for HAV did not detect this new HAV genotype, knowledge of its existence may ameliorate potential significant epidemiological and research implications in laboratories globally.

Possible Concomitant Aggressive NK Cell Leukemia and EBV-positive T-cell lymphoma; Using the online beta version of WHO-HAEM5 and videoconferencing software to make diagnoses accessible in an emerging economy

BackgroundUsing the World Health Organization Classification 5th edition (beta version online; WHO-HAEM5bv) in emerging economies is key to global healthcare equity. Although there may be ongoing updates, hesitancy in accepting and reporting these diagnoses in publication conflicts with the WHO’s commitment to global accessibility. Aggressive NK cell leukemia (ANKL) and systemic EBV-positive T-cell lymphoma of childhood (SEBVTCL) with CD4-positive immunophenotype are both rare entities, are most described in Asians and East Asians, are associated with prior systemic chronic active EBV disease (CAEBV), and presentation with Hemophagocytic Lymphohistiocytosis (HLH). Recognizing and diagnosing any one of these entities requires not only training and experience in hematopathology, but good cooperation between clinical physicians and all areas of the laboratory. We describe a 30-year-old woman who presented to a Vietnam hospital and was rapidly diagnosed with ANKL, SEBVTCL, and HLH using WHO-HAEM5bv essential criteria, aided by expert consultation from a United States (US) board certified hematopathologist in real-time using video conferencing software.MethodsZoom™ videoconferencing software; Immunohistochemistry; flow cytometric immunophenotyping; polymerase chain reaction (PCR), Next Generation Sequencing (NGS).ResultsAt the time of hospital admission, automated complete blood count (CBC) with differential count showed slight anemia, slight lymphocytosis, and moderate thrombocytopenia. HIV serology was negative. Whole blood PCR for EBV was positive showing 98,000 copies/ml. A lymph node biopsy revealed histology and immunohistochemistry consistent with the online beta version WHO-HAEM5 classification of SEBVTCL arising in CAEBV. Blood and bone marrow studies performed for staging revealed no histologic or immunohistochemical evidence of T-cell lymphoma in the bone marrow core, however, atypical blood smear lymphocyte morphology and blood immunophenotyping by flow cytometry were consistent with WHO-HAEM5 classification of ANKL. NGS revealed no evidence of genetic variant(s) associated with HLH in Vietnam. All laboratory studies were performed at Blood Transfusion Hematology Hospital (BTHH) in Ho Chi Minh City Vietnam.ConclusionAlthough Vietnam, an emerging economy, currently lacks the laboratory infrastructure to more rigorously confirm a rare synchronous presentation of two distinct EBV-driven T/NK cell neoplasms, these two concomitant diagnoses were made using only laboratory techniques available in Vietnam with the help of WHO-HAEM5bv and real-time video consultation by a US hematopathologist.

Proteomics identification of overexpressed serum proteins in dogs with Babesiacanis infection.

Canine babesiosis, caused by the protozoan parasite Babesia canis, is characterized by clinical manifestations, including hemolytic anemia, thrombocytopenia, multiple organ failure, and may result in death. This disease is detected using conventional blood smears, which are time-consuming and have low sensitivity. This study aimed to investigate a more rapid and sensitive method for detecting B. canis infection in dogs by examining the expressed serum protein profiles using proteomics. We collected six sera samples from three healthy and three B. canis-infected dogs diagnosed using blood smear and polymerase chain reaction. We analyzed the proteins using two-dimensional gel electrophoresis. The candidate spots from the gel were subjected to protein identification using a nano-liquid chromatography system coupled to an ion-trap mass spectrometer equipped with an electrospray ionization nano-sprayer. We found that 10 protein spots were overexpressed in the serum samples from infected dogs compared with healthy dogs, which corresponded to three proteins: serotransferrin, serotransferrin isoforms X1, and hemopexin. Furthermore, analysis of the protein-protein interaction network confirmed that they strongly interacted with each other. This study suggests that high levels of serotransferrin and hemopexin are related to B. canis infection, making these proteins potential candidates for the development of diagnostic molecules or vaccines.

Clinical, Haemato-biochemical Evaluation and Comparison of Polymerase chain reaction with Conventional Blood Smear Examination for the Detection of Babesia gibsoni Infection in Dogs

The objective of the present study was to detect Babesia gibsoni infection in dogs which are presented to Veterinary Clinical Complex, Rajiv Gandhi Institute of Veterinary Education and Research, Puducherry. A total of 85 dogs with clinical signs such as inappetence, fever, enlarged lymph nodes and tick infestation were screened. The diagnostic efficacy of polymerase chain reaction (PCR) technique has been compared with blood smear examination to detect B. gibsoni infection in dogs. By blood smear and PCR examination, six (7.1%) dogs and 28 (42.4%) dogs were positive for B. gibsoni infection respectively. Clinical examination of dogs with B. gibsoni showed tick infestation, anorexia, dullness, pyrexia, pale/congested conjunctival mucus membrane, tachypnoea and swollen lymph nodes. In addition, few dogs had icterus and haemoglobinuria. Laboratory examination of blood samples of B. gibsoni dogs revealed significant reduction in haemoglobin, PCV, erythrocyte count, platelet count and serum albumin levels. Significant increase in levels of ALP, globulin, bilirubin, BUN and creatinine were recorded.

Development of a multiplex reverse transcription qPCR assay for identification of saliva, blood, and semen

ABSTRACT The identification of body fluids can provide important information for case processing in forensic investigations. Accurate identification through messenger RNA (mRNA) is effective, especially in cases of sexual assault, where body fluids from one or more contributors are mixed. In this study, a saliva, blood, and semen simultaneous identification reverse transcription-quantitative polymerase chain reaction (SBS RT-qPCR) assay using mRNA from a DNA/RNA co-extraction method was developed. The body fluid-specific mRNA markers histatin 3 (HTN3), haemoglobin subunit beta (HBB), and protamine 1 (PRM1) were selected for identification of saliva, blood, and sperm cells, respectively. The assay accurately identified saliva, blood, and semen without cross-reaction with 10 body fluids. The limits of detection (LoD) were 1/102-, 1/105-, and 1/103-fold dilutions of total RNA in saliva, blood, and semen, respectively. Furthermore, the assay accurately identified trace amounts of body fluids in mixed samples at a 58:1:1 ratio. Finally, it was possible to analyse the origin of body fluids by obtaining an STR profile using the DNA/RNA co-extraction method. The SBS RT-qPCR assay was able to simultaneously identify each body fluid with efficiency in sample consumption. Therefore, it is advisable to use the DNA/RNA co-extraction method to identify mixed body fluids in sexual assault cases.