Risk factors associated with the presence of Mycobacterium bovis in macroscopic lesions suspected as being caused by bovine tuberculosis detected in slaughterhouses

Mycobacterium bovis is a bacterium belonging to the Mycobacterium tuberculosis complex that causes tuberculosis in cattle and in other domestic and wild animals, as well as in humans. Disease control measures are carried out by slaughtering animals tested positive in the intradermal tuberculinization test and sanitation of their original living spaces, in addition to epidemiological surveillance carried out through the sanitary inspection of bovine carcasses in slaughterhouses. In the latter, official inspection services collect samples from macroscopic lesions suspected of bovine tuberculosis, which are then sent for laboratory analysis. Knowledge concerning the variables associated with the occurrence of M. bovis can aid in decision-making regarding control and disease eradication efforts. In this context, the aim of this study was to identify the risk factors for a positive M. bovis diagnosis in suspected bovine tuberculosis lesions obtained during epidemiological surveillance activities in the state of Mato Grosso, Brazil. A total of 105 suspicious lesions were analyzed using the Nested Polymerase Chain Reaction (nested q-PCR) method, of which 14 (13.33%) tested positive for M. bovis. Univariate and bivariate statistical analyses indicated that the variable “animal slaughter” was the only risk factor presenting statistical significance associated with the diagnosis of M. bovis (p < 0.05), demonstrating that macroscopic lesions suspected as being caused by bovine tuberculosis from animals with an in vivo diagnosis were 2.82 - fold more likely to result in a positive M. bovis diagnosis by molecular tests.

Chagas Disease in Pregnant Women from Endemic Regions Attending the Hospital General de Mexico, Mexico City

Trypanosoma cruzi infection leads to Chagas disease (CD), a neglected tropical infection of significant public health importance in South and Central America and other, non-endemic, countries. Pregnant women and their children are of particular importance to screen as T. cruzi can be transmitted vertically. The objective of this study was to screen for T. cruzi infection among pregnant women from endemic areas seen at the Hospital General de Mexico for prenatal care, so that they and their children may be quickly connected to CD treatment. Pregnant women were recruited through the hospital prenatal clinic and screened for T. cruzi infection using a series of serological and molecular tests. Of 150 screened patients, mean age 26.8 (SD 6.4), 30 (20.0%) were positive by at least one diagnostic test. Of these, only nine (6%) were positive as determined by PCR. Diagnosis of chronic CD is difficult in endemic places like Mexico due to the limitations of current commercially available diagnostic tests. Further evaluation of diagnostic performance of various assays could improve current CD diagnostic algorithms and proper care management in these regions. Genetic variability in the parasite may also play a role in the differing assay performances seen in this study, and this may be a valuable avenue of further research.

Drug susceptibility testing of Mycobacterium tuberculosis using next generation sequencing and Mykrobe software

Introduction. Mycobacterium tuberculosis is the causative agent of tuberculosis. Drug susceptibility testing is performed by phenotypic and molecular tests. Commonly used for phenotypic drug susceptibility testing is the automated BACTEC system in a liquid culture medium. Drug susceptibility by line probe molecular tests was introduced almost 15 years ago. Recently whole genome sequencing (WGS) analysis of M. tuberculosis strains demonstrated that genotyping of drug-resistance could be accurately performed. Several software tools were developed.Our study aimed to perform whole-genome sequencing on phenotypically confirmed multi-drug resistant (MDR) M. tuberculosis strains, to identify drug-resistant mutations and to compare whole-genome sequencing profiles with line probe assay and phenotypic results.Materials and methods. We performed analysis on 34 MDR M. tuberculosis Bulgarian strains. Phenotypic drug susceptibility testing was performed on the BACTEC system. For molecular testing of drug susceptibility to first- and second-line tuberculostatics, we applied line probe assay Geno Type MTBDR plus v.1.0 и Geno Type MTBDR sl v.1.0. Sequencing was performed on MiSeq. Generated FASTQ files were analyzed for known drugresistant mutations with the software platform Mykrobe v.0.8.1.Results. All three methods — phenotypic analysis using the BACTEC system, genetic analysis of strains applying the Geno Type test and Mykrobe software gave comparable sensitivity/resistance results for the studied strains. All phenotypically proven rifampicin and isoniazid-resistant strains were 100% confirmed using Mykrobe software. The C-15T mutation is a marker for isoniazid resistance in strains of the SIT41 spoligotype. We observed a 75% (21/28) agreement between BACTEC and Mykrobe for ethambutol resistance. Phenotypically, 87% (n = 27) of the strains are resistant to streptomycin, but only 59% (n = 19) are proven by Mykrobe software. Comparing phenotypic and genotypic resistance to ofloxacin, amikacin and kanamycin, we observed 100% coincidence of results.Conclusions. Whole-genome sequencing approach is relatively expensive and laborious but useful for detailed analysis such as epidemiological genotyping and molecular drug susceptibility testing.

Konya İli Böğürtlen, Ahududu ve Kuşburnu Yetiştiriciliğinde Potansiyel Bir Tehdit: Ateş Yanıklığı Hastalığı

In the present study, totally 49 samples, which showed the symptoms of leaf and shoot blight and cankers with brown discoloration of necrotic tissues on mature branches, were collected from 22 districts and areas of Konya Province between 2017 and 2019. Presence rate of E. amylovora in collected samples, showing symptoms of the disease, from the province was determined to be 40% for blackberry and raspberry and 33% rosehip for rosehip in three years. Bacteria consistently isolated from the diseased tissues were identified on the basis of biochemical, physiological, and molecular tests, comparing with a reference strain of E. amylovora, isolated from blackberry (Kbb 371). Twenty seven representative bacterial strains were gram-negative, rod-shaped, mucoid, fermentative, positive for levan formation and acetoin production, no growth at 36°C, positive for gelatin hydrolysis, and negative for esculin hydrolysis, indole, urease, catalase, oxidase, arginine dehydrolase, reduction of nitrate, acid production from lactose, and inositol. All strains induced a hypersensitive response in tobacco (Nicotiana tobacum cv. White Burley) 24 h after inoculation with a 108 CFU ml-1 bacterial suspension in sterile distilled water. The strains were identified as E. amylovora using the species-specific primers set A/B (1), which amplified a 1-kb DNA fragment in PCR, and the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method. In order to fulfill the Koch postulates, pathogenicity test was confirmed by injecting bacterial suspensions of 108 CFU ml-1 in sterile distilled water into the shoot tips of 3-year-old blackberry R. fruticosus cv. Chester, raspberry R. idaeus cv. Heritage and rosehip R. canina. All tests were repeated three times. The bacterium was re-isolated from inoculated plants and identified as E. amylovora. Phytosanitary measures are needed to prevent any further spread of the bacterium as potential inoculum sources to new blackberry, raspberry and rosehip growing areas.

Identification of equine herpesvirus 8 in donkey abortion: a case report

Background Equine herpesvirus-8 (EHV-8) is one of the most economically significant viruses that infect mammals of the genus Equus worldwide, which cause severe respiratory diseases and abortion in horses. However, there is no report of abortion caused by EHV-8 in donkeys. Case presentation The present case report is about a 4-year-old donkey having an abortion and showing a serious respiratory issue on the 296th day of pregnancy. Bacteriological and molecular tests were used to screen possible bacterial/viral pathogens to detect the etiological agent. Salmonella abortus equi, EHV-1, EHV-4, and EAV were all negative in the current study. EHV-8, on the other hand, was the only agent that was isolated and identified. Conclusions This was for the first time that EHV-8 had been isolated from a donkey in China. EHV-8 infection can cause abortion in donkeys; therefore, veterinarians and breeders should be aware of it.

Amplificação isotérmica mediada por loop para detecção de patógenos de plantas

Disease control is crucial to minimize potential losses in agriculture and thereby maintain high crop yield. However, for its effectiveness, the pathogen must be detected early and correctly in the production fields. Different methods of diagnosis can be used, from those based on symptoms to molecular tests. Loop-mediated isothermal amplification (LAMP) is a molecular technique that has been widely used in several biological fields, due to the ease with which it can be applied. The reaction can be carried out in a single thermal condition, due to the use of Bst DNA polymerase, isolated from the bacterium Bacillus stearothermophilus, which has high displacement activity. LAMP is a highly exponential amplification method that produces the target DNA in amounts 109 -1010 times between 45 and 60 minutes at 60-65°C. Its advantages are the visualization of results directly with the naked eye and the fact that it does not need sophisticated equipment for its application. In phytopathology, the technique has been gaining prominence in the detection of fungi, viruses, bacteria, nematodes and phytoplasmas, as well as in the monitoring of fungicide-resistant fungi. LAMP can benefit agriculture so that early, accurate and sensitive diagnostics can be carried out in the fields of cultivation and minimize losses caused by diseases. In this review, we present and discuss LAMP tests, developed for plant pathogens detection, which can be useful for researchers who wish to use the technique in their research area

Contemporary Molecular Analyses of Malignant Tumors for Precision Treatment and the Implication in Oral Squamous Cell Carcinoma

New molecular tests and methods, in addition to morphology-based diagnosis, are widely used as a new standard of care in many tumors. “One-size-fits-all medicine” is now shifting to precision medicine. This review is intended to discuss the key steps toward to development of precision medicine and its implication in oral squamous cell carcinoma. The challenges and opportunities of precision medicine in oral cancer will be sequentially discussed based on the four steps of precision medicine: identification/detection, diagnosis, treatment and monitoring.

Sensitivity and Specificity of Serology, Histopathology, and Molecular Tests in the Detection of Leptospirosis from Slaughtered Cattle in Indonesia

Leptospira spp. is a pathogenic bacteria that causes leptospirosis in humans and cattle. The World Health Organization (WHO) recommends the microscopic agglutination test (MAT) as the laboratory gold standard in the detection of leptospirosis. However, the limitation of MAT triggers the laboratory technicians to develop alternative laboratory tests against leptospirosis. The current study aimed to compare the sensitivity and specificity of histopathology special stain using modified Gram staining (MGS) and molecular test using reverse transcriptase-polymerase chain reaction (RT-PCR), compared to the MAT for Leptospira spp. detection in cattle. This study used a total of 38 serum and 38 kidney samples from the cattle slaughtered in the Sidoarjo slaughterhouse, Indonesia. The collected serum samples were tested against MAT and RT-PCR. The kidneys were processed for histopathology using MGS. The result indicated that 16 (42.10%) of the tested samples were positive against MAT, 6 (15.78%) were positive against MGS, and 18 (47.36%) were positive against RT-PCR. The RT-PCR indicated better sensitivity and lower specificity, compared to MAT and MGS. The findings revealed that the RT-PCR is an appropriate laboratory test for detecting cattle leptospirosis with better sensitivity and specificity. Therefore, this method can be suggested to substitute MAT and overcome its limitations.

Evaluation of the BioFire Gastrointestinal Panel to Detect Diarrheal Pathogens in Pediatric Patients

Infectious diarrhea is a global pediatric health concern; therefore, rapid and accurate detection of enteropathogens is vital. We evaluated the BioFire® FilmArray® Gastrointestinal (GI) Panel with that of comparator laboratory tests. Stool samples of pediatric patients with diarrhea were prospectively collected and tested. As a comparator method for bacteria, culture, conventional PCR for diarrheagenic E. coli, and Allplex GI-Bacteria(I) Assay were tested. For discrepancy analysis, BD MAX Enteric Bacterial Panel was used. As a comparator method for virus, BD MAX Enteric Virus Panel and immunochromatography was used and Allplex GI-Virus Assay was used for discrepancy analysis. The “true positive” was defined as culture-positive and/or positive results from more than two molecular tests. Of the 184 stool samples tested, 93 (50.5%) were true positive for 128 pathogens, and 31 (16.9%) were positive for multiple pathogens. The BioFire GI Panel detected 123 pathogens in 90 of samples. The BioFire GI Panel demonstrated a sensitivity of 100% for 12 targets and a specificity of >95% for 16 targets. The overall positive rate and multiple pathogen rate among patients in the group without underlying diseases were significantly higher than those in the group with hematologic disease (57.0% vs. 28.6% (p = 0.001) and 20.4% vs. 4.8% (p = 0.02), respectively). The BioFire GI Panel provides comprehensive results within 2 h and may be useful for the rapid identification of enteropathogens.

Dengue 2 serotype and yellow fever coinfection

Case Presentation. Arboviruses primarily consist of RNA, which favours greater genetic plasticity, with a higher frequency of mutations that allow the virus to adapt to different hosts. The initial symptomatology is nonspecific, in that the patient can present fever, myalgia, arthralgia, rash and headache. This makes a clinical diagnosis using laboratory tests difficult and time-consuming. In Brazil, the main arboviruses involved in epidemics belong to the family Flaviviridae. The patient in this case is from the municipality of São Bernardo do Campo, an area endemic for arboviruses. He presented symptoms of fever, myalgia and headache. Results. The multiplex assay for arboviruses detected genetic material from the dengue 2 and yellow fever viruses. Conclusion. This result confirms the importance of molecular tests showing high sensitivity and specificity that can assist clinical diagnosis, particularly in endemic areas during periods of outbreak for other arboviruses, like the epidemiological picture in Brazil in 2018, when significant co-circulation of dengue virus and yellow fever virus occurred. The presence of co-circulating arboviruses increases the chance of coinfection and demonstrates the importance of differential diagnosis.