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In order to prevent spread of COVID-19, World Health Organization (WHO) has specified that measures such as cleaning hands regularly with alcohol-based hand sanitizer or washing with soap and water, avoiding touching nose, eyes, mouth and social distancing should be followed by people. Another important measure for containing spread is by Testing–Testing and Testing. By testing, patients affected by COVID-19 can be isolated and further spread from them can be prevented. For testing, clinical specimen inactivation should be performed in a bio-safety level-3 (BSL-3) laboratory and after this step, routine testing can be performed in a bio-safety level-2 laboratory. Based on the infection and death rates in the current pandemic situation, few high-level bio-safety facilities are required to handle this deadly virus associated with potential exposure risks. It would take a long time, a large staff and many resources to construct a new fixed bio-safety facility, thus delaying the prevention and control of the pandemic. Therefore, a mobile BSL-3 laboratory with both bio-safety and flexibility is urgently needed to manage the pandemic and emergent public health incidents. To meet this requirement, the author has designed a mobile virology research and diagnostic laboratory with inputs from ESIC Medical College, Hyderabad, identified two companies which specialize in fabrication of shelters and containment facilities, realized and installed it within 15 days of time. This state-of-the-art, one of its kind laboratory was inaugurated by Hon’ble Raksha Mantri on 23 April, 2020. The MVRDL developed as per WHO and ICMR bio-safety standard (BSL-3) can be used:To conduct real-time reverse transcription polymerase chain reaction (rRT-PCR) test for diagnosing COVID-19. Possibility to test up to 2000 samples per day.For virus culturing for drug screening, convalescent plasma-derived therapy.To aid in the development of vaccine and development of diagnostic kits.

The ongoing pandemic of SARS-CoV-2 calls for rapid and cost-effective methods to accurately identify infected individuals. The vast majority of patient samples is assessed for viral RNA presence by RT-qPCR. Our biomedical research institute, in collaboration between partner hospitals and an accredited clinical diagnostic laboratory, established a diagnostic testing pipeline that has reported on more than 40,000 RT-qPCR results since its commencement at the beginning of April 2020. However, due to ongoing demand and competition for critical resources, alternative testing strategies were sought. In this work, we present a clinically-validated standard operating procedure (SOP) for high-throughput SARS-CoV-2 detection by RT-LAMP in 25 minutes that is robust, reliable, repeatable, sensitive, specific, and inexpensive.

The SARS-CoV-2 pandemic has resulted in unprecedented challenges to veterinary diagnostic laboratories. These challenges include partial or complete shutdowns, interrupted courier services, disruptions in workflow and diagnostic testing, new physical distancing practices, protocol development or enhancement for handling samples from high-risk or susceptible species, and fulfilling requirements for pre-test permission approval from state and federal veterinary agencies, all of which have been implemented to prevent or minimize exposure and transmission of SARS-CoV-2 locally or regionally. As in people, SARS-CoV-2 infects animals through direct animal-to-animal contact and aerosol transmission between animals. Humans can also infect pets or other animals in their care and, although human-to-human transmission is the main route of viral spread in people, infected animals and specimens of their bodily fluids or tissues are a potential source of infection for veterinarians and technical or laboratory personnel that are handling them. In this perspective, we discuss how SARS-CoV-2 has necessitated rapid changes in laboratory operation to minimize zoonotic risk to personnel and to implement tests for identifying the virus in animals. The pandemic has highlighted the adaptability and quick response of veterinary diagnosticians to an emerging infectious disease and their critical role in maintaining animal health, while synergizing with and protecting human public health.

Introduction: We report on the first documented cluster of Coronavirus Disease 2019 cases amongst diagnostic laboratory staff and outline some of the initial and ongoing steps that are being implemented to manage and prevent the spread of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in our laboratory Case presentation: On 24 April 2020, three staff members of a tertiary diagnostic laboratory in Groote Schuur Hospital, Cape Town, South Africa, tested positive for SARS-CoV-2 Within seven days, a further nine cases were identified, which suggested an outbreak and prompted a full investigation Management and outcome: A multifaceted strategic approach was adopted to halt the spread of SARS-CoV-2 in our laboratory Interventions focused on simultaneously establishing appropriate risk mitigation and stratification strategies through the upscaling of infection prevention and control measures, whilst minimising disruption to service delivery Conclusion: Laboratory Coronavirus Disease 2019 outbreaks have the potential to cripple a laboratory’s testing capacity Contingency planning and risk assessments should occur early, and interventions should be modified according to each laboratory’s available resources and infrastructure FAU - Opperman, Christoffel J

When South Florida became a hotspot for COVID-19 disease in March 2020, we faced an urgent need to develop test capability to detect SARS-CoV-2 infection. We assembled a transdisciplinary team of knowledgeable and dedicated physicians, scientists, technologists and administrators, who rapidly built a multi-platform, PCR- and serology-based detection program, established drive-thru facilities and drafted and implemented guidelines that enabled efficient testing of our patients and employees. This process was extremely complex, due to the limited availability of needed reagents, but outreach to our research scientists and to multiple diagnostic laboratory companies and government officials enabled us to implement both FDA authorized and laboratory developed testing (LDT)-based testing protocols. We analyzed our workforce needs and created teams of appropriately skilled and certified workers, to safely process patient samples and conduct SARS-CoV-2 testing and contact tracing. We initiated smart test ordering, interfaced all testing platforms with our electronic medical record, and went from zero testing capacity, to testing hundreds of healthcare workers and patients daily, within three weeks. We believe our experience can inform the efforts of others, when faced with a crisis situation.

Accessory tragus is a congenital malformation of part of the external ear, commonly reported in humans. Clinically, it is a benign, cutaneous mass located anywhere between the tragus of the ear and the angle of the mouth, along the migratory path of the first branchial arch. An accessory tragus was diagnosed in an otherwise healthy six-month-old male castrated American pit bull terrier that had a haired, pedunculated cutaneous mass on the left maxillary region from birth. Histologically, the mass was a polypoid extension of histologically normal haired skin with a central core of subcutaneous adipose tissue and well-differentiated elastic cartilage. To the authors' knowledge, this is the first report of this lesion in a non-human species. Retrospective examination of records from 2008 to 2018 at the Washington Animal Disease Diagnostic Laboratory failed to identify any other case. As accessory tragus in humans is commonly linked with other congenital anomalies and syndromes, recognition of this lesion in animals may aid in early discovery of other congenital defects and inform adequate excision of the lesion to prevent chondrodermatitis.

Background One major challenge for detecting the virus that causes COVID-19 is commercial SARS-CoV-2 testing kit or reagent availability. To allow every laboratory or hospital access to an in-house assay, we developed two low-cost SARS-CoV-2 detection assay protocols using in-house primers and reagents/equipment on hand in most biology or diagnostic laboratories: a SYBR Green–based RT-PCR and PCR assays. RNA extraction has also become a major bottleneck due to limited supplies and the required labor. Thus, we validated alternative RNA extraction protocols. Methods SARS-CoV-2 genome sequences deposited into the GISAID database were retrieved to design and synthesize in-house primers. Forty patient samples were collected by nasopharyngeal swab, coded, and used to develop and validate the assay protocols. Both assays used TRIzol and heat-processing techniques to extract RNA from patient samples and to inactivate the virus; thus, testing was conducted in a conventional biosafety level 2 laboratory. Results The sensitivity and specificity of the primers were evaluated using samples previously confirmed positive for SARS-CoV-2. The positive amplicons were sequenced to confirm the results. The assay protocols were developed, and the specificity of each PCR product was confirmed using melting curve analyses. The most accurate heat-processing technique for primers with short amplicon lengths was 95 °C for 15 mins. Of 40 samples, both the SYBR Green–based quantitative RT-PCR assay and the PCR assay detected SARS-CoV-2 target genes in 28 samples, with no false-positive or false-negative results. These findings were 100% concordant with those of the diagnostic laboratory that tested the same samples using a Rotor-Gene PCR cycler with an Altona Diagnostics SARS-CoV-2 kit (R2=0.889). Conclusions These approaches are reliable, repeatable, specific, sensitive, simple, and low-cost tools for the detection of SARS-CoV-2 in a conventional biosafety level 2 laboratory, offering alternative approaches when commercial kits are unavailable or cost ineffective.

A pandemia do Coronavírus 2 (SARS-CoV-2), iniciada na China, e ainda em curso no Brasil e mundo afora, já registra mais de 4,0 milhões de casos confirmados e mais de 280 mil óbitos após 4 meses de dispersão do vírus. É no momento o patógeno de maior prioridade a ser tratado, por ter alta transmissibilidade, associado à graves doenças e mortalidade, assim como, ausência de medidas profiláticas e terapêuticas eficazes, limitados conhecimentos da epidemiologia, imunidade e patogênese humana. Para acompanhar à crescente demanda dos serviços clínicos e diagnósticos, é necessário aprimorar a biossegurança e expandir a capacidades dos laboratórios e profissionais. O artigo destaca a classe de risco e os consensos dos requisitos mínimos de biossegurança para atividades em laboratórios clínicos e diagnósticos com o SARS-COV-2, assim como, a prevenção de riscos biológicos aos profissionais, evitando disseminação acidental às famílias e à comunidade.

The emergence of the novel coronavirus SARS-CoV-2 has led to a pandemic infecting more than two million people worldwide in less than four months, posing a major threat to healthcare systems. This is compounded by the shortage of available tests causing numerous healthcare workers to unnecessarily self-isolate. We provide a roadmap instructing how a research institute can be repurposed in the midst of this crisis, in collaboration with partner hospitals and an established diagnostic laboratory, harnessing existing expertise in virus handling, robotics, PCR, and data science to derive a rapid, high throughput diagnostic testing pipeline for detecting SARS-CoV-2 in patients with suspected COVID-19. The pipeline is used to detect SARS-CoV-2 from combined nose-throat swabs and endotracheal secretions/ bronchoalveolar lavage fluid. Notably, it relies on a series of in-house buffers for virus inactivation and the extraction of viral RNA, thereby reducing the dependency on commercial suppliers at times of global shortage. We use a commercial RT-PCR assay, from BGI, and results are reported with a bespoke online web application that integrates with the healthcare digital system. This strategy facilitates the remote reporting of thousands of samples a day with a turnaround time of under 24 hours, universally applicable to laboratories worldwide.

Treatment and control of bovine respiratory disease (BRD) is predicated on the use of two categories of antimicrobials, namely bacteriostatic drugs that inhibit bacterial growth and replication (STATIC), and bactericidal drugs that kill bacteria in in vitro culture systems (CIDAL). Recently, we reported that initial BRD treatment with a STATIC antimicrobial followed by retreatment with a CIDAL antimicrobial was associated with a higher frequency of multidrug-resistant bacteria isolated from field cases of BRD submitted to a veterinary diagnostic laboratory. The present study was conducted to test the hypothesis that calves administered the same class of antimicrobial for first and second BRD treatment (i.e., CIDAL-CIDAL or STATIC-STATIC) would have improved health and performance outcomes at the feedlot compared to calves that received a different antimicrobial class for retreatment (i.e., STATIC-CIDAL or CIDAL-STATIC). The association between antimicrobial treatments and health, performance, and carcass quality outcomes were determined by a retrospective analysis of 4,252 BRD treatment records from a commercial feedlot operation collected from 2001 to 2005. Data were compared using generalized linear mixed statistical models that included gender, season, and arrival weight as covariates. The mean (±SE) probability of BRD cases identified as requiring four or more treatments compared to three treatments was greater in calves that received STATIC-CIDAL (73.58 ± 2.38%) or STATIC-STATIC (71.32 ± 2.52%) first and second antimicrobial treatments compared to calves receiving CIDAL-CIDAL (50.35 ± 3.46%) first and second treatments (P < 0.001). Calves receiving CIDAL-CIDAL first and second treatments also had an increased average daily gain (1.11 ± 0.03 kg/d) compared to calves receiving STATIC-CIDAL (0.95 ± 0.03 kg/d) and STATIC-STATIC (0.84 ± 0.02 kg/d) treatments (P < 0.001). Furthermore, CIDAL-CIDAL-treated calves had a higher probability of a choice quality grade at slaughter (36.44 ± 4.80%) compared to STATIC-CIDAL calves (28.09 ± 3.88%) (P = 0.037). There was no effect of antimicrobial treatment combination on BRD mortality (P = 0.855) or yield grade (P = 0.240) outcomes. These observations suggest that consideration should be given to antimicrobial pharmacodynamics when selecting drugs for retreatment of BRD. These findings have implications for developing BRD treatment protocols that address both post-treatment production and antimicrobial stewardship concerns.

Objectives:Clinical diagnostics in sudden onset disasters have historically been limited. We set out to design, implement, and evaluate a mobile diagnostic laboratory accompanying a type 2 emergency medical team (EMT) field hospital.Methods:Available diagnostic platforms were reviewed and selected against in field need. Platforms included HemoCue301/WBC DIFF, i-STAT, BIOFIRE FILMARRAY multiplex rt-PCR, Olympus BX53 microscopy, ABO/Rh grouping, and specific rapid diagnostic tests. This equipment was trialed in Katherine, Australia, and Dili, Timor-Leste.Results:During the initial deployment, an evaluation of FilmArray tests was successful using blood culture identification, gastrointestinal, and respiratory panels. HemoCue301 (n = 20) hemoglobin values were compared on Sysmex XN 550 (r = 0.94). HemoCue WBC DIFF had some variation, dependent on the cell, when compared with Sysmex XN 550 (r = 0.88-0.16). i-STAT showed nonsignificant differences against Vitros 250. Further evaluation of FilmArray in Dili, Timor-Leste, diagnosed 117 pathogens on 168 FilmArray pouches, including 25 separate organisms on blood culture and 4 separate cerebrospinal fluid pathogens.Conclusion:This mobile laboratory represents a major advance in sudden onset disaster. Setup of the service was quick (< 24 hr) and transport to site rapid. Future deployment in fragmented health systems after sudden onset disasters with EMT2 will now allow broader diagnostic capability.

Background Notwithstanding the efforts of direct-acting antivirals (DAAs) for the treatment of chronically infected hepatitis C virus (HCV) patients, concerns exist regarding the emergence of resistance-associated substitutions (RAS) related to therapy failure. Sanger sequencing is still the reference technique used for the detection of RAS and it detects viral variants present up to 15%, meaning that minority variants are undetectable, using this technique. To date, many studies are focused on the analysis of the impact of HCV low variants using next-generation sequencing (NGS) techniques, but the importance of these minority variants is still debated, and importantly, a common data analysis method is still not defined. Methods Serum samples from four patients failing DAAs therapy were collected at baseline and failure, and amplification of NS3, NS5A and NS5B genes was performed on each sample. The genes amplified were sequenced using Sanger and NGS Illumina sequencing and the data generated were analyzed with different approaches. Three different NGS data analysis methods, two homemade in silico pipeline and one commercially available certified user-friendly software, were used to detect low-level variants. Results The NGS approach allowed to infer also very-low level virus variants. Moreover, data processing allowed to generate high accuracy data which results in reduction in the error rates for each single sequence polymorphism. The results improved the detection of low-level viral variants in the HCV quasispecies of the analyzed patients, and in one patient a low-level RAS related to treatment failure was identified. Importantly, the results obtained from only two out of the three data analysis strategies were in complete agreement in terms of both detection and frequency of RAS. Conclusions These results highlight the need to find a robust NGS data analysis method to standardize NGS results for a better comprehension of the clinical role of low-level HCV variants. Based on the extreme importance of data analysis approaches for wet-data interpretation, a detailed description of the used pipelines and further standardization of the in silico analysis could allow increasing diagnostic laboratory networking to unleash true potentials of NGS.

Herpes simplex keratitis (HSK) is a common, potentially blinding condition characterised by recurrent infections of the cornea, seen by both general ophthalmologists and corneal specialists. Successful treatment of recurrences reduces disease duration, prevents progressive corneal scaring leading to vision loss and reduces risk of further recurrences. In this review we discuss the relative advantages of the diagnostic laboratory investigations including polymerase chain reaction, viral culture and fluorescence-based immunohistochemistry. We review treatment strategies in selected aspects of HSK and discuss the management options in cases not responding to treatment.摘要: 单纯疱疹病毒性角膜炎(herpes simplex keratitis, HSK) 是一种常见、潜在的致盲性疾病, 其特征是角膜反复感染, 普通眼科医生和角膜专家都可见到该病。成功治疗其反复发作可缩短病程, 防止进行性角膜瘢痕形成所导致的视力丧失以及降低进一步复发的风险。在这篇综述中, 我们讨论了实验室诊断包括聚合酶链反应、病毒培养及基于荧光的免疫组化的相对优势。我们对HSK在某些方面的治疗策略进行了回顾, 以及针对治疗无效病例的处理方案。.

Background Since the epidemiology of canine and feline dermatophytosis might evolve in response to chronological, sociological and ecological factors, the authors studied the occurrence of dermatophyte pathogens over 27 years subsequent to the last major UK survey.\ud \ud Methods Dermatophyte culture submission records from dogs and cats to the Royal Veterinary College Diagnostic Laboratory in England between 1991 and 2017 were reviewed. Samples were routinely cultured aerobically at 26°C for up to four weeks on Sabouraud’s dextrose agar containing cycloheximide and chloramphenicol; dermatophytes were identified using conventional phenotypic methods.\ud \ud Results Proportional isolation from cats (15.9 per cent of 1389) exceeded that of dogs (8.1 per cent of 2193) (P

An 8-mo-old, crossbred, heifer calf was presented to the Heeke Animal Disease Diagnostic Laboratory with a history of ataxia and altered mentation. Grossly, the liver was diffusely yellow-orange, turgid, and exuded watery, thin blood on cut section. The cortex and medulla in both kidneys were diffusely and markedly dark brown to black. The urinary bladder was filled with dark red urine. Histologically, centrilobular hepatocellular degeneration was observed, but these sections lacked necrosis. In the kidney, numerous cortical tubules contained intraluminal bright eosinophilic fluid and red-orange granular casts that stained positive for hemoglobin with the Dunn–Thompson method. The gross and histologic lesions supported a high level of suspicion for copper toxicosis. Feed and water samples from the farm were submitted for mineral analysis. The copper concentration in the feed was 118 mg/kg, and the molybdenum concentration was 0.9 mg/kg. Chronic copper toxicosis is rarely reported in cattle. The gross lesions in our case are a departure from, although similar to, previously reported cases, including lack of histologic hepatocellular necrosis. Collectively, gross and histologic lesions were compatible with copper toxicosis in this calf, and copper concentrations in the feed samples suggest a feed-mixing error.

Whole exome sequencing (WES)-based assays undergo rigorous validation before being implemented in diagnostic laboratories. This validation process generates experimental evidence that allows laboratories to predict the performance of the intended assay. The NA12878 Genome in a Bottle (GIAB) HapMap reference sample is commonly used for validation in diagnostic laboratories. We investigated what data points should be taken into consideration when validating WES-based assays using the GIAB reference in a diagnostic setting. We delineate specific factors that require special consideration and identify OMIM genes associated with diseases that may ‘bypass’ validation. Four replicates of the NA12878 sample were sequenced at the CHEO Genetics Diagnostic Laboratory on a NextSeq 500; the data were analyzed using the bcbio_nexgen v1.1.2 pipeline. The hap.py validation engine, Real Time Genomics vcfeval tool, and high confidence (HC) variant calls in HC regions available for the GIAB sample were used to validate the obtained variant calls. The same validation process was then used to evaluate variant calls obtained for the same sample by two other clinical diagnostic laboratories. We showed that variant calls in NA12878 can be confidently measured only in the regions that intersect between the GIAB HC regions and the target regions of exome capture. Of the 4139 (as of October 2019) OMIM genes associated with a phenotype and having a known molecular basis of disease, 84 were fully outside of the GIAB HC regions and many of the remaining OMIM genes were only partially covered by the HC regions. A significant proportion of variants identified in the NA12878 sample outside of the HC regions have unknown (UNK) status due to the absence of HC reference alleles. Verification of such calls is possible either by an alternative truth set or by orthogonal testing. Similarly, many variants outside of exome capture regions, if not accounted for, will be deemed false negatives due to insufficient probe coverage. Our results demonstrate the importance of the intersection between genomic regions of interest, capture regions, and the high confidence regions. If not considered, false and ambiguous variant calls could have a negative impact on diagnostic accuracy of the intended WES-based diagnostic assay and increase the need for confirmatory testing. To enable laboratories to identify ‘problematic’ regions and optimize validation efforts, we have made our VCF and BED files available in UCSC Genome Browser: NA12878 WES Benchmark. Relevant genes and genome annotations are evolving, we implemented a general purpose algorithm to cross-reference OMIM genes with the genomic regions of interest that can be applied to capture genes/regions outside HC regions (see repository of data material section).

More than twelve blood group systems have been described in dogs, but little is known about their distribution frequencies within breed populations. Here, we report on an extensive typing survey carried out using available reagents and either established or new clinical kits in purebred dogs from Germany. Leftover anticoagulated blood samples were examined using an immunochromatographic strip method for DEA 1, a gel column technique for Dal and Kai 1/2, and new card agglutination tests for DEA 4 and DEA 5 (which were partially compared with the gel column technique). Monoclonal antibodies were used for DEA 1 and Kai 1/2 typing, and polyclonal antibodies were used for all other types. Among the 206 dogs, 59.2% were DEA 1+, 100% DEA 4+, 9% (Card)/11% (Gel) DEA 5+, 89.3% Dal+, 96.6% Kai 1+, and 2.9% Kai 2+. None of the dogs were Kai 1+/2+, and only one was Kai 1-/2-. Dal- dogs were found in several breeds. Erythrocytes from most DEA 1+ dogs bound strongly on the strips. The agglutination reactions for DEA 5 on the new card tests were generally less than those on the gel column. The blood group pattern DEA 4+, DEA 5-, Dal+, Kai 1+/2- and either DEA 1+ or DEA 1- was found among 80% of the dogs. In this first extensive blood typing survey of purebred dogs from Europe, the proportions of positive and negative blood types were similar to those found in the United States and, for DEA 1, were also similar to those from other European countries, with considerable breed variation in blood types. The newer typing techniques seem to work well and will likely be useful for detecting and preventing specific blood type incompatibilities in the clinic.

Background - The PHE Public Health Laboratory, Birmingham at Heartlands hospital has an annual turn-over in excess of 750,000 specimens, 41,000 of which are processed using real-time PCR in the molecular section. In 2017, due to the age of existing instrumentation, the laboratory began a programme of replacement. Methods - During 2017-19 the entire molecular service was replaced with a new high-throughput automated Roche FLOW system, consisting of a primary sample handler (PSU), two MagNA Pure 96 instruments, a PCR set-up unit (PSU), three 384-well LightCycler 480 II instruments and overarching FLOW software responsible for automated data handling, specimen tracking and workflow between the instruments. Results - The bulk of the in-house developed assay repertoire, representing 34 viral, bacterial and fungal targets was re-optimised in 19 multiplexes. These were fully validated against existing molecular tests and were introduced into routine diagnostic service in June 2018. The FLOW software was successfully interfaced with the LIMS system, allowing rapid two-way communication of test requests and results. Over the following ten months, the service was expanded with an additional seven targets. The new system and assays were audited by UKAS in April 2019 and achieved accreditation to ISO:15189:2012 in August 2019. Conclusion – The Roche FLOW system was successfully validated and introduced into routine diagnostic service and now offers significant improvements in assay performance, sample throughput and turnaround time. Details of the validation process, post-implementation modifications, trouble-shooting and lessons learnt will be of value to other diagnostic laboratories considering the introduction of this technology.

Frontotemporal dementia (FTD) is a heterogeneous neurodegenerative disorder characterized by frontal and temporal lobe atrophy, typically manifesting with behavioural or language impairment. Because of its heterogeneity and lack of available diagnostic laboratory tests there can be a substantial delay in diagnosis. Cell-free, circulating, microRNAs are increasingly investigated as biomarkers for neurodegeneration, but their value in FTD is not yet established. In this study, we investigate microRNAs as biomarkers for FTD diagnosis. We performed next generation small RNA sequencing on cell-free plasma from 52 FTD cases and 21 controls. The analysis revealed the diagnostic importance of 20 circulating endogenous miRNAs in distinguishing FTD cases from controls. The study was repeated in an independent second cohort of 117 FTD cases and 35 controls. The combinatorial microRNA signature from the first cohort, precisely diagnosed FTD samples in a second cohort. To further increase the generalizability of the prediction, we implemented machine learning techniques in a merged dataset of the two cohorts, which resulted in a comparable or improved classification precision with a smaller panel of miRNA classifiers. In addition, there are intriguing molecular commonalities with cell free miRNA signature in ALS, a motor neuron disease that resides on a pathological continuum with FTD. However, the signature that describes the ALS-FTD spectrum is not shared with blood miRNA profiles of patients with multiple sclerosis. Thus, microRNAs are promising FTD biomarkers that might enable earlier detection of FTD and improve accurate identification of patients for clinical trials

While the semen analysis remains as the cornerstone for the evaluation of male infertility, there have been significant recent advances in diagnostic laboratory techniques, assisted-reproductive technologies, and proteomics. The evolving changes in diagnostic criteria and therapeutic interventions has necessitated a more comprehensive and methodological assessment of infertile men. The aim of this chapter is to discuss modern laboratory exams and techniques available to successfully evaluate and treat male factor infertility.

The objective of this work was to conduct a retrospective study of causes of diarrhea in calves in southern Rio Grande do Sul between 1980 and 2014. All necropsy protocols and materials received in the Regional Diagnostic Laboratory of the Veterinary School of the Federal University of Pelotas were reviewed. Ninety-four cases of calves with diarrhea were diagnosed, of these, 44.7% were caused by parasitic infections and 34.0% by bacterial infections. Parasites as cause of diarrhea was the most important diagnostic in southern Rio Grande do Sul and affected calves from three to 12 months of age. The occurrence of diarrhea in animals even infants may be associated with environmental factors and the form of management that these animals are kept. Bacterial diseases are also important causes of diarrhea in calves up to three months of age in the region.

Objective. This study is aimed at investigating if point-of-care testing for HbA1c (POCT-HbA1c) using the HemoCue® HbA1c 501 system could be an alternative method for diabetes screening and monitoring to replace the HbA1c measurement in a standard diagnostic laboratory. Design. This was a cross-sectional study to assess the agreement between POCT and a standard laboratory measurement method for determining the level of HbA1c. Setting and Participants. In total, 108 participants were recruited to participate in this study, consisting of 61 diabetics and 47 nondiabetics. The diabetic group comprised 37 females and 24 males, diagnosed with type 2 diabetes mellitus (DM) and undergoing diabetes treatment at several community health care centres in Bandung, West Java. The nondiabetic group consisted of 15 female and 32 male patients of several community health care centres and healthy volunteers. Sample Collection and Analysis. A venous blood sample was taken for routine HbA1c analysis by the diagnostic laboratory method. For the POCT-HbA1c, a blood sample was taken from the fingertip at the same time and analysed with the HemoCue® HbA1c 501 system. Outcome Measures. The HbA1c results of both methods were compared and analysed with a Bland-Altman agreement plot. The sensitivity and specificity of the POCT-HbA1c data were also compared with those of the standard diagnostic results. Results. Based on the Bland-Altman plot, the HbA1c level for 100 out of 108 (92.59%) subjects analysed by the POCT-HbA1c was within the range of the 95% limit of agreement. Compared with the standard diagnostic assay, the sensitivity of the POCT-HbA1c was 97.83% and its specificity was 77.42%. Conclusions. The high sensitivity and accuracy of POCT-HbA1c indicate that it is a potential method for diabetes screening and monitoring to replace the routine diagnostic laboratory HbA1c measurement, especially when a rapid result is required.

The SARS-CoV-2 pandemic has resulted in shortages of both critical reagents for nucleic acid purification and highly trained staff as supply chains are strained by high demand, public health measures and frequent quarantining and isolation of staff. This created the need for alternate workflows with limited reliance on specialised reagents, equipment and staff. We present here the validation and implementation of such a workflow for preparing samples for downstream SARS-CoV-2 RT-PCR using liquid handling robots. The rapid sample preparation technique evaluated, which included sample centrifugation and heating prior to RT-PCR, showed a 97.37% (95% CI: 92.55-99.28%) positive percent agreement and 97.30% (95% CI: 90.67-99.52%) negative percent agreement compared to nucleic acid purification-based testing. This method was subsequently adopted as the primary sample preparation method in the Groote Schuur Hospital Virology Diagnostic Laboratory in Cape Town, South Africa.

Fluorescent in situ hybridization (FISH) techniques can be used to identify a range of chromosome abnormalities that are clinically significant in many cancers. Multicolor FISH can be used to identify multiple targets, which can be simultaneously detected in individual cells using digital imaging microscopy. In an era of precision medicine there is a requirement to make a precise diagnosis and to have a molecular classification of the tumor that can guide therapy. Cancer genomics is now regarded as a sub-specialism in pathology and genomic testing needs to be robustly integrated into the routine diagnostic practice.The FISH techniques described in this chapter have been developed over many years in a busy hematopathology diagnostic laboratory. We describe robust in-house methods for both liquid samples (blood and bone marrow mainly) and formalin-fixed paraffin-embedded (FFPE) tissue biopsies that allow for large numbers of slides to be set up in batches. The techniques described are for interphase cells in tissues where metaphase chromosome techniques are generally not applicable. Some of the FISH tests need to be carried out as an "out-of-hours" emergency test to make a critical diagnosis while others provide prognostic information and are used to guide downstream patient management.

Until mass vaccination befalls, control of the new betacoronavirus-associated severe acute respiratory syndrome pandemic (SARS-CoV-2) is based on decreasing virus circulation by social distancing and blocking transmission foci after diagnosis. Globally adopted SARS-CoV-2 diagnostic criteria embrace viral RNA detection by quantitative reverse-transcription polymerase chain reaction (qRT-PCR) on nasopharynx secretions, which requires healthcare facilities and specialized personnel for sample collection. To develop an alternative protocol, hydrophilic cotton as the material and saliva as the source for biological sample collection in qRT-PCR/RT-endpoint-PCR SARS-CoV-2 diagnostic methods prepared with local consumables were evaluated using 99 archived nasopharynx samples previously diagnosed as positive for SARS-CoV-2 and 111 prospective saliva samples pared with nasopharynx samples from patients attending the local reference ABC Medical School diagnostic laboratory. The kappa agreement coefficient between the SARS-CoV-2 qRT-PCR and RT-endpoint-PCR was k = 0.97 (95% CI 0.92-1.00) and k = 0.90 (95% CI 0.81-0.99), respectively, on SARS-CoV-2-positive archived samples, with the initial qRT-PCR CT under 25. The agreement coefficient of the SARS-CoV-2 alternative saliva diagnostic protocol, when used to test the paired nasopharynx samples, was k = 0.79 (95% CI 0.56-1,00). These data support that the SARS-CoV-2 diagnostic assay based on self-collected saliva on cotton represents an alternative protocol for mass diagnosis and epidemiological studies in low-income regions.

The molecular diagnostic laboratories in developing countries are progressively improving and attaining international accreditation. Hence, understanding and implementing the concept of quality are the needs of the hour. Total quality management system (TQMS) in a diagnostic laboratory is an integrated program involving all laboratory staff aiming for integration, consistency, and increase in efficiency. A comprehensive understanding of such a system is unfortunately only through practice. Our endeavors in establishment of TQMS for laboratory accreditation has helped us gain comprehensive understanding of the process and have compelled us to share the knowledge with the community engaged in such diagnostics. A number of accreditation bodies have put forth guidelines for laboratories seeking accreditation; however, such documents can be overwhelming at times leading to confusion and difficulty in implementation. We aim to present to the reader a lucid script of basic reasoning behind such accreditation guidelines in conjunction with thorough and practical generalized workflows for TQMS implementation. The goal of TQMS is to provide the best possible care for patients; and to achieve top-notch quality, there should be a continuous drive for improvement. The clinical vignettes presented in this paper illustrate the value of applying select ISO 15189:2012 recommendations in a molecular diagnostic laboratory to efficiently ensure quality through TQMS and a basic understanding of nuances for its practical implementation. Implementation of TQMS in a diagnostic setting enables seamless flow of laboratory results and instills confidence in clinicians regarding disease interpretation and treatment.

Background Influenza A Virus (IAV) causes respiratory disease in swine and is a zoonotic pathogen. Uncontrolled IAV in swine herds not only affects animal health, it also impacts production through increased costs associated with treatment and prevention efforts. The Iowa State University Veterinary Diagnostic Laboratory (ISU VDL) diagnoses influenza respiratory disease in swine and provides epidemiological analyses on samples submitted by veterinarians. Description To assess the incidence of IAV in swine and inform stakeholders, the ISU FLUture website was developed as an interactive visualization tool that allows the exploration of the ISU VDL swine IAV aggregate data in the clinical diagnostic database. The information associated with diagnostic cases has varying levels of completeness and is anonymous, but minimally contains: sample collection date, specimen type, and IAV subtype. Many IAV positive samples are sequenced, and in these cases, the hemagglutinin (HA) sequence and genetic classification are completed. These data are collected and presented on ISU FLUture in near real-time, and more than 6,000 IAV positive diagnostic cases and their epidemiological and evolutionary information since 2003 are presented to date. The database and web interface provides rapid and unique insight into the trends of IAV derived from both large- and small-scale swine farms across the United States of America. Conclusion ISU FLUture provides a suite of web-based tools to allow stakeholders to search for trends and correlations in IAV case metadata in swine from the ISU VDL. Since the database infrastructure is updated in near real-time and is integrated within a high-volume veterinary diagnostic laboratory, earlier detection is now possible for emerging IAV in swine that subsequently cause vaccination and control challenges. The access to real-time swine IAV data provides a link with the national USDA swine IAV surveillance system and allows veterinarians to make objective decisions regarding the management and control of IAV in swine. The website is publicly accessible at http://influenza.cvm.iastate.edu. Electronic supplementary material The online version of this article (10.1186/s12859-018-2408-7) contains supplementary material, which is available to authorized users.

To determine the epidemiology of Candida auris in South Africa, we reviewed data from public- and private-sector diagnostic laboratories that reported confirmed and probable cases of invasive disease and colonization for October 2012–November 2016. We defined a case as a first isolation of C. auris from any specimen from a person of any age admitted to any healthcare facility in South Africa. We defined probable cases as cases where the diagnostic laboratory had used a nonconfirmatory biochemical identification method and C. haemulonii was cultured. We analyzed 1,692 cases; 93% were from private-sector healthcare facilities, and 92% of cases from known locations were from Gauteng Province. Of cases with available data, 29% were invasive infections. The number of cases increased from 18 (October 2012–November 2013) to 861 (October 2015–November 2016). Our results show a large increase in C. auris cases during the study period, centered on private hospitals in Gauteng Province.

The analysis of statistical data on detection of onychomycosis pathogens in the clinical diagnostic laboratory of the regional dermatovenerologic dispensary of the city of Dnipro from 2014 to 2018 and the Center of Laboratory Medicine «Vis-Medic» for the first half of 2018. The clinical manifestations of onychomycosis and principles of their therapy are considered. Special attention was paid to the use of lacquer with cyclopirox (Cyclogen) in local therapy. In view of the particularities of the chemical structure, effects and treatment results, Cyclogen can be considered as a qualitative alternative to solutions with antifungal drug., Был проведен анализ статистических данных о выявляемости возбудителей онихомикозов в клинико-диагностической лаборатории областного кожно-венерологического диспансера г. Днепра с 2014 по 2018 г. и Центре лабораторной медицины «Вис-Медик» за 1-е полугодие 2018 г. Рассмотрены клинические проявления онихомикозов и принципы их терапии. Уделено особое внимание применению в местной терапии лака с циклопироксом («Цикложен»). Учитывая особенности химического строения, действие и результаты лечения, «Цикложен» может рассматриваться как качественная альтернатива растворам с противогрибковым препаратом., Був проведений аналіз статистичних даних про виявлення збудників оніхомікозів у клініко-діагностичній лабораторії обласного шкірно-венерологічного диспансеру м. Дніпра з 2014 до 2018 р. і Центрі лабораторної медицини «Віс-Медик» за 1-е півріччя 2018 р. Розглянуто клінічні вияви оніхомікозів і принципи їх терапії. Приділено особ­ливу увагу застосуванню в місцевій терапії лаку з циклопіроксом («Цикложен»). З огляду на особливості хімічної будови, дії і результатів лікування, «Цикложен» можна розглядати як якісну альтернативу розчинам з протигрибковим препаратом.

BackgroundCopper associated hepatitis (CAH) has been increasingly recognized in dogs, and speculation exists that hereditary defects in copper metabolism have been exacerbated by increased environmental copper exposure. However, no broad epidemiological investigations have been performed to investigate quantitative hepatic copper concentrations ([Cu]H ) over time in both dogs that are (predisposed breed [PB]), and are not (non-predisposed breed [NPB]), considered at-risk for CAH.ObjectivesTo investigate [Cu]H in dogs and explore temporal, demographic, and histologic associations spanning 34 years.Animals546 archived liver specimens.MethodsRetrospective study. Searches of the Michigan State University Veterinary Diagnostic Laboratory database identified dogs that had undergone hepatic histopathologic assessment. Cases with archived tissue were reviewed and classified by breed, time period, and presence or absence of hepatitis. Inductively coupled plasma mass spectrometry was used to determine [Cu]H .ResultsIn time period 2009-2015, median [Cu]H were 101 μg/g and 313 μg/g greater than median [Cu]H in time period 1982-1988 for NPB and PB dogs, respectively (P < .001 for both comparisons). The proportion of dogs with [CU]H > 300 μg/g increased in NPB (28% to 49%) and PB dogs (48% to 71%) during these periods (P = .002 for both comparisons). Median [Cu]H in dogs with hepatitis increased 3-fold over time in both NPB (P = .004) and PB populations (P < .001).Conclusions and clinical importanceThe frequent recognition of CAH in recent years is likely due to the observed increases in [Cu]H over time. Importantly, effects are not limited to PB dogs.

This article describes the need for, stratifies the complexity of, and proposes detailed lists of training competencies for diagnostic laboratory personnel using quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) for patient care. Although quantitative LC-MS/MS is evolving toward greater automation with less need for technical expertise, gaps remain in resources for training and assessment.

This chapter presents the limitations of diagnostic testing for drugs. Diagnostic laboratory investigations for drugs of misuse have a number of functions, including diagnosis, monitoring the impact of treatment, monitoring compliance with treatment in coerced treatment systems, such as those involving court-enforced drug orders, and motivational reinforcement of people’s progress. The chapter explains the varying time windows of detection of sampling types and substances. The tests that are available can detect substances that are used or misused in urine, blood, sweat, oral fluid and hair. The window of detection varies with the substance, the method of its use, and the sampling medium. The use of urine samples for testing for substances raises issues of privacy and dignity, as collecting an observed urine sample is the only way to ensure absolutely that the sample is unadulterated and from the particular patient.

Companion animal abuse is an issue that concerns not only veterinarians and law enforcement agencies but also society in general. Animals that die under suspicious or violent circumstances should be submitted to reference laboratories for a postmortem examination by veterinary forensic pathologists trained to recognize animal abuse. Nevertheless, the low notification rate of such cases in Spain may explain the country's lag in the development of veterinary forensics and the limited information available on animal abuse epidemiology in comparison with other countries. By sharing information among the entities involved in recognizing animal abuse, we can better address these issues, thus improving veterinary forensics in Spain (and elsewhere). In this study, we analysed the cause and manner of death of 96 dog and cat carcasses suspected of animal abuse. These cases were submitted to our diagnostic laboratory for forensic postmortem examination by public agencies and animal protection centres. To our knowledge, this is the first study to focus on the postmortem analysis of forensic cases of suspected dog and cat abuse in Spain. On the basis of gross and histopathological findings, we distinguished between natural and non-natural abuse-related deaths, classifying the latter. We confirmed that most of the dog deaths were related with abuse, though the suspected abuse and the cause of death did not always coincide. In contrast, cause of death was determined to be natural in many of the cat suspected abuse cases. The most frequent non-natural abuse-related cause of death in dogs was blunt force trauma (n=24, 43.64%), followed by firearm injuries (n=10, 18.18%), asphyxia (n=5, 9.09%), heatstroke (n=3, 5.45%), starvation (n=2, 3.64%), bite injury (n=1, 1.82%), and sharp force trauma (n=1, 1.82%). In cats, the most common cause of death was blunt force trauma (n=9, 21.95%), followed by firearm injuries (n=3, 7.32%) and bite injury (n=2, 4.88%). The main goal of our study is to share our results with the scientific community to advance the field of veterinary forensics in Spain, which will lead to more successful prosecutions by law enforcement agencies. Finally, we highlight that veterinarians who can accurately recognize signs of animal abuse may be able to better respond to cases of companion animal cruelty, which in turn, may prevent possible escalation to interpersonal violence.

Johne's disease has historically been considered a disease of dairy cattle with little impact on beef herds in the United States. Over the past several years, a substantial increase in the number ofJohne's positive submissions from Midwestern beef operations has been observed at the Iowa State University Veterinary Diagnostic Laboratory, suggesting an increase in the prevalence of disease in the cow-calf industry. Thus, there is an urgent need to better understand the current knowledge ofbeefveterinarians related to Johne's disease and assess the impact of the disease in the cow-calf industry., American Association of Bovine Practitioners Proceedings of the Annual Conference, 2018

The histopathological image classification is a vivid application for medical diagnosis and neural network has been successful in the image classification task. However, finding the optimal values of the neural network is still a challenging task. To accomplish the same, this paper considers a two-layer neural network which is optimized through intelligent gravitational search algorithm. Further, the optimized two-layer neural network is applied for the histopathological tissue classification into healthy and inflamed. The proposed method is validated on the publicly available tissue dataset, namely Animal Diagnostic Laboratory (ADL). The experimental results firm the better performance of the proposed method against state-of-the-art methods in terms of seven performance measures, namely recall, specificity, precision, false negative rate (FNR), accuracy, F1-score, and G-mean.

Objetivo: promover o debate sobre a importância do laboratório no apoio à saúde pública e aos serviços de saúde nos países de língua portuguesa, numa perspetiva global e de uma só saúde. Métodos: revisão de literatura, análise dos documentos preparatórios das comunicações do 5º Congresso Nacional de Medicina Tropical, apresentações realizadas pelos oradores de Cabo Verde, Angola e Portugal e o debate que se seguiu. Resultados: identificaram-se os desafios que deverão ser ultrapassados de forma a criar uma rede de laboratórios de diagnóstico que possa estimular o acesso universal das populações aos cuidados de saúde, tendo-se proposto medidas e estratégias com base na experiência dos sistemas de saúde dos diferentes países de língua portuguesa. Reviu-se o contributo dos Laboratórios do IHMT/NOVA para esta estratégia. Conclusão: não há acesso universal a cuidados de saúde de qualidade sem serviços de diagnóstico laboratorial de qualidade. Assim, deve-se promover uma correta, eficaz e sustentável inter-relação entre laboratórios de referência nacionais, hospitalares e privados e ainda laboratórios de investigação, todos cooperando em sintonia com os serviços de saúde de modo a garantir os melhores cuidados de saúde., Anais do Instituto de Higiene e Medicina Tropical, v. 18 (2019): Políticas, Sistemas e Serviços de Saúde - 5º Congresso Nacional de Medicina Tropical

The Veterinary Diagnostic Laboratory has the great role in diagnosing medical testing for infectious agents, toxins, and other causes of disease in animal diagnostic samples which submitted by veterinary practitioners serving animal owners, public health offi cials, wildlife management, and scientists with research projects. This Veterinary Diagnostic Laboratory guidance focused on the discipline of special veterinary faculty like pathology, clinical pathology, microbiology, virology, immunology, parasitology, and serology.

Bovine anaplasmosis (BA) is tick-borne disease of cattle caused by Anaplasma marginale and it remains an economically important disease in the United States (U.S.). We have anecdotal information that Veterinary Feed Directive prescriptions in Kentucky (KY) are written most often for treatment and prevention of BA. However, there are no recent prevalence estimates of this disease in KY. Thus, this study was aimed at determining the seroprevalence of and factors associated with BA in KY. Data were obtained from an active slaughter survey (n = 232) performed between May and July 2013 as well as from reviewing The University of Kentucky Veterinary Diagnostic Laboratory (UKVDL) records of specimens submitted for BA testing from 2002–2012 (n = 2,573). With competitive ELISA, the apparent prevalence of BA in KY was 10.78% (95% CI: 7.41–15.42%) and 11.58% (95% CI: 10.31–12.98%) for the slaughter survey and laboratory records, respectively. Whereas the estimated true prevalence was 9.44% (95% CI: 5.65–14.48%) and 10.3% (95% CI: 8.92–11.8%), respectively. From the laboratory records, factors associated with positive BA results were age, breed, whether specimens were submitted singularly or as a group, year and quarter of the year the specimens were submitted. The odds of the outcome were 5 times as high when cattle were adults (vs juvenile) and almost 4 times as high when specimens were submitted singularly (vs group). In comparison to Holstein breed, the odds of the outcome were 3.5 and 2.5 times higher in Angus and mixed breeds, respectively. The odds of a diagnosis of BA varied in an undulating pattern by year of sample submission. When compared to 2011, the odds of a diagnosis of BA was approximately 3 times as high in 2005, 2008, and 2009 and approximately 5 times as high in 2004, 2006, and 2012. In comparison to the duration from January to March, the odds of the outcome were almost 20 times as high from July to September but 10 times as high from October to December durations. Counties with specimen submissions for BA testing had a significantly greater cattle population and number of cattle farms than counties without specimen submissions. Future prevention and control measures for BA should target these factors and should be weighted more on counties with higher cattle population. Furthermore, current records from the UKVDL appear sufficient for the surveillance of BA in KY.

Розглянуто актуальні питання управління якістю медичних послуг і втілення їх в практичну діяльність. Серйозні проблеми, пов’язані із якістю результатів досліджень, створює багато об’єктивних і суб’єктивних чинників. Для усунення проблемних чинників розпочато розроблення національних стандартів у галузі лабораторної медицини з використанням міжнародного досвіду розроблення стандартів, що регламентують вимоги до організації діяльності клініко-дігностичних лабораторій та засобів лабораторного аналізу. Гострою проблемою лабораторних досліджень є забезпечення точності результатів. У Європейському співтоваристві давно існують вимоги щодо забезпечення метрологічної простежуваності калібраторів і контрольних матеріалів та метрологічного забезпечення лабораторного устаткування, а також методик досліджень. Забезпечення якості лабораторних досліджень є однією з центральних проблем сучасної лабораторної медицини. Основною вимогою для таких досліджень є надання клієнтам надійної інформації. Це, передусім, визначає вимогу щодо якості клінічних лабораторних досліджень клінічних і діагностичних лабораторій, що забезпечує правильне та своєчасне призначення аналізу пацієнту, виконане на належному аналітичному рівні з необхідною інформацією для його інтерпретації. Тільки за умови чіткої організації та якісного проведення лабораторних досліджень можна очікувати, що кожен результат, відображений у авторизованому звіті, лікар зможе використати для прийняття діагностичних рішень або рішень, які змінюють схему лікування. Одним зі способів визначення компетентності клініко-діагностичних лабораторій та якості їх досліджень є проведення незалежної оцінки якості. Лабораторна медицина надає 70–80 % інформації, необхідної для прийняття рішень щодо діагностики, лікування та профілактики захворювань. Надання достовірної інформації клініцистам – важлива вимога до якості клінічних лабораторних досліджень клініко-діагностичних лабораторій, яка забезпечує правильне і своєчасне призначення аналізу пацієнтові. Тільки за умови якісного виконання лабораторного дослідження можна розраховувати на правильне прийняття діагностичних рішень і належне лікування.

Introduction: Widal test has been used extensively for the sero-diagnosis of Enteric fever in India, however, its accuracy and reliability are debatable. We studied widal testing and widal positivity rates in the entire city of Ahmedabad for the diagnosis of Enteric Fever. Methods We screened all 1700 possible diagnostic laboratory facilities, in Ahmedabad, in the public and private sector. We performed telephonic surveys for the initial filtering of facilities that could be conducting widal testing. It was followed by physical visits to probable facilities to confirm testing methods and preservation of reports of widal testing. We followed a systematic process for screening and selection of 23 laboratories, which conducted widal tests and had reliable data. While 14 laboratories refused to share data, data provided by three of them were inappropriate and couldn’t be used. We finally analyzed data from four large public hospitals, one private trust hospital and one corporate laboratory for variable periods in a span of 15 years (2000 – 2015). Result: The Widal testing rate was found to be 8.7% and widal positivity as 12.5% in a sample of 1.2 million clinically suspected in-patients. In 15 years, the private hospital had admitted 1/10th as many cases as all the public hospitals together. However, the widal testing and positivity rates were similar in both. We observed a lower proportion of widal positivity among children below 12 years and a disproportionate, but insignificant, gender distribution of widal positivity. Conclusion: This study indicates that the widal test, which is meant to be an initial screening test, is widely used in the city. We propose linkage of testing and reporting of widal with other more reliable and accurate tests such as Typhidot and blood culture in order to strengthen our knowledge of enteric fever epidemiology in India.

Introduction: Hepatitis C virus (HCV) shows great structural variability. Based on genome sequencing and phylogenetical analysis, 7 types and 67 subtypes can be differentiated with varying geographical distribution. It is very important to determine the HCV type/subtype prior to starting direct antiviral therapy (DAA), which has been available since 2014, because the type, dose and optimal length of medication depends on these. Aim: In Hungary, the treatment of chronic HCV patients started in 1992 with the relevant special diagnostic tests being carried out in our Molecular Diagnostic Laboratory. Determination of the nucleotide sequence of the Hungarian HCV1b NS5A/PKR-BR region and the type and subtype distribution of Hungarian patients have already been carried out. The current summary discusses the results of 6092 chronic HCV patients (175 serotypes, 5917 genotypes) based on age, gender, regions and genotype distribution changes over the period between 1996 and 2017. Method: Serotyping (1996–1999). Genotyping: hybridization (2000–2016), real-time PCR (2016–; Cobas 4800 HCV GT). Results: Genotype distribution: GT1a: 5.6%; GT1b: 84.6%; GT1a + 1b: 5.1%; GT2: 0.1%; GT3: 1.8%; GT4: 0.1%; mixed: 1.6%; GT1 (non-differentiated subtype): 1,1%. Women/men ratio: 52%/48%. The most common age category is 50–60 years (37% of all cases). There was no genotype asymmetry among the four Hungarian regions and Budapest. Over time, the prevalence of genotype 3 increased from 1.6% to 2.8% and the number of patients under the age of 40 doubled. Conclusion: There have been no substantial changes in the HCV type/subtype distribution in Hungary over the past 20 years, 1b remaining the most common. The introduction of real-time PCR method for genotyping has resulted in a major quality improvement including only a few mixed subtype results leading to more efficient drug selection. Orv Hetil. 2018; 159(Suppl 2): 2–8.

Recommendations for bovine mycoplasma culture CO 2 concentrations are varied and were not empirically derived. The objective of this study was to determine whether the growth measures of bovine mycoplasma isolates differed when incubated in CO 2 concentrations of 10 or 5% or in candle jars (2.7 ± 0.2% CO 2 ). Growth of Mycoplasma bovis (n = 22), Mycoplasma californicum (n = 18), and other Mycoplasma spp. (n = 10) laboratory isolates was evaluated. Isolate suspensions were standardized to approximately 10 8 cfu/mL and serially diluted in pasteurized whole milk to achieve test suspensions of 10 2 and 10 6 cfu/mL. One hundred microliters of each test dilution was spread in duplicate onto the surface of a modified Hayflick's agar plate. Colony growth was enumerated on d 3, 5, and 7 of incubation. A mixed linear model included the fixed effects of CO 2 treatment (2.7, 5, or 10%), species, day (3, 5, or 7), and their interactions, with total colony counts as the dependent variable. Carbon dioxide concentration did not significantly affect overall mycoplasma growth differences, but differences between species and day were present. Colony counts (log 10 cfu/mL) of M. bovis were 2.6- and 1.6-fold greater than M. californicum and other Mycoplasma spp., respectively. Growth at 7 d of incubation was greater than d 3 and 5 for all species. These findings were confirmed using field isolates (n = 98) from a commercial veterinary diagnostic laboratory. Binary growth responses (yes/no) of the field isolates were not different between CO 2 treatments but did differ between species and day of incubation. On average, 57% of all field isolates were detected by 3 d of incubation compared with 93% on d 7. These results suggest that the range of suitable CO 2 culture conditions and incubation times for the common mastitis-causing Mycoplasma spp. may be broader than currently recommended.

Autoantibodies are helpful tools not only for the diagnosis and the classification of systemic autoimmune rheumatic diseases (SARD) but also for sub-grouping patients and/or for monitoring disease activity or specific tissue/organ damage. Consequently, the role of the diagnostic laboratory in the management of SARD is becoming more and more important. The advent of new techniques raised the need of updating and harmonizing our use/interpretation of the assays. We discuss in this opinion paper some of these issues. Indirect immunofluorescence (IIF) was originally suggested as the reference technique for anti-nuclear antibody (ANA) detection as previous solid phase assays (SPA) displayed lower sensitivity. The new available SPA are now offering better results and can represent alternative or even complementary diagnostic tools for ANA detection. The improved sensitivity of SPA technology is also changing our interpretation of the results for other types of autoantibody assays, but we need updating their calibration and new reference materials are going to be obtained in order to harmonize the assays. There is growing evidence that the identification of autoantibody combinations or profiles is helpful in improving diagnosis, patients’ subgrouping and predictivity for disease evolution in the field of SARD. We report some explanatory examples to support the idea to make the use of these autoantibody profiles more and more popular. The technological evolution of the autoimmune assays is going to change our routine diagnostic laboratory tests for SARD and validation of new algorithms is needed in order to harmonize our approach to the issue.

Veterinary diagnostic laboratories (VDLs) play a significant role in the prevention and mitigation of endemic animal diseases and serve an important role in surveillance of, and the response to, outbreaks of transboundary and emerging animal diseases. They also allow for business continuity in livestock operations and help improve human health. Despite these critical societal roles, there is no academic literature on the economic impact of VDLs. We present a case study on the economic impact of the Iowa State University Veterinary Diagnostic Laboratory (ISUVDL). We use economic contribution analysis coupled with a stakeholder survey to estimate the impact. Results suggest that the ISUVDL is responsible for $2,162.46 million in direct output, $2,832.45 million in total output, $1,158.19 million in total value added, and $31.79 million in state taxes in normal years. In an animal health emergency this increases to $8,446.21 million in direct output, $11,063.06 million in total output, $4,523.70 million in total value added, and $124.15 million in state taxes. The ISUVDL receives $4 million annually as a direct state government appropriation for operating purposes. The $31.79 million in state taxes in normal years and the $124.15 million in state taxes in an animal health emergency equates to a 795% and 3104% return on investment, respectively. Estimates of the economic impact of the ISUVDL provide information to scientists, administrators, and policymakers regarding the efficacy and return on investment of VDLs.

Detection of circulating antigen of Dirofilaria immitis has been a mainstay of identifying heartworm infection in clinical practice for the past three decades. Several validated commercial antigen tests have very good sensitivity, specificity, and positive predictive values, especially when used in patients for which heartworm infection is likely. In some dogs and cats infected with heartworm, antigen may not be available for detection although present in the patient sample; heat pretreatment of these samples reveals the antigen, changing the false negative to positive. This phenomenon was documented in the literature in the 1980s but subsequently overlooked by the heartworm research community for many years. In this review, we provide a summary of the current understanding of the role of heat reversal in diagnosing heartworm infection. This additional diagnostic step is most important for patients in which heartworm infection is likely, such as dogs or cats in an endemic area with an inconsistent history of heartworm preventive use, or dogs with a prior diagnosis of heartworm infection that were recently treated. To illustrate the concept, we share a summary of results from canine samples tested at the state veterinary diagnostic laboratory in Oklahoma, USA in 2017 by modified Knott test and by commercial antigen test before and after heat treatment of samples; in this sample set, heat treatment changed all D. immitis microfilaria-positive but antigen-negative samples to antigen-positive. Pet dogs with a history of consistent preventive use are unlikely to become positive with heat pretreatment; for that reason, routine pretreatment of all samples tested in a veterinary practice is not recommended. We also review known causes of false negative and false positive results on heartworm antigen tests that, although uncommon, can complicate accurate diagnosis in individual patients. Together, this review provides a primer to aid understanding of strategies that can enhance accurate diagnosis of heartworm infection in veterinary practice and clinical research.

The diagnostic laboratory is essential to patient care and to the achievement of health equity. Through the development of quality laboratories in settings burdened by poverty and weak health systems, Partners In Health has demonstrated the critical contributions of clinical laboratories to the care of patients with HIV, tuberculosis, and cancer, among other conditions. The lessons learned through the organization's experience include the importance of well-trained and well-supported staff; reliable access to supplies, reagents, and diagnostic equipment; adequate facilities to provide diagnostic services; the integration of laboratories into networks of care; and accompaniment of the public health sector.

Background Li-Fraumeni syndrome (LFS) has traditionally been identified by single-gene testing (SGT) of TP53 triggered by clinical criteria, but the widespread use of multigene panel tests (MGPTs) has upended this paradigm. We sought to compare the personal and family cancer histories of TP53-positive result (TP53+) carriers who were identified by either MGPT or SGT. Methods Of 44 310 individuals who underwent testing of TP53 in a single clinical diagnostic laboratory between 2010 and 2014, 44 086 (40 885 MGPT and 3201 SGT) met study eligibility criteria. Personal cancer histories were available for 38 938 subjects. The frequency of germline TP53 results and various phenotypic manifestations were compared according to test type. All statistical tests were two-sided. Results MGPT TP53+ individuals (n = 126) had an older median age at first cancer than SGT TP53+ carriers (n = 96; women: median = 36 vs 28 years, P < .001; and men: median = 40 vs 15 years, P = .004). The median age of breast cancer diagnosis was 40 years in MGPT TP53+ women vs 33 years in SGT TP53+ women (P < .001). In both cohorts, childhood and LFS core cancers, and for women, multiple primary cancers (not multiple breast tumors), were associated with TP53+ results. Established LFS testing criteria were less often met by MGPT TP53+ individuals. Conclusions MGPT TP53+ individuals differ in phenotype from those ascertained through SGT and are notably older at cancer diagnosis and less likely to meet LFS clinical criteria. These findings suggest that LFS may have a greater phenotypic spectrum than previously appreciated. This has implications for the counseling of MGPT TP53+ individuals. Prospective follow-up of these individuals and families is needed to re-evaluate cancer risks.