Prevalence of Pulmonary Mycotic Lesions and Antifungal Susceptibility Pattern of Fungi Isolated from Nigerian Trade Cattle

Pulmonary mycosis is a major cause of deaths in many livestock herds, especially in immune-compromised animals. A total of 138 lung samples (6mm in size) were collected from cattle slaughtered for meat at Nsukka abattoir. Swab sticks were dipped into each lung sample and streaked on freshly prepared Sabouraud Dextrose Agar (SDA) for fungal isolation. Isolated fungi were identified using morphological and microscopic features. Disc diffusion method was used to test the antifungal susceptibility pattern of a randomly selected isolates with voriconazole (1μg), fluconazole (25μg) and methanol extract of Garcinia kola, using disc diffusion method. The lung samples were examined for lesions using histopathological procedures. Out of the 138 lung samples, 23 had fungal growths while 115 had no growth of any sort. A total of 34 fungi were isolated as pure cultures. Aspergillus spp. had the highest frequency of isolation, 44.92%, followed by Penicillium spp. 11.76%, Trichophyton spp. 5.88%, Conidiobolus and Sporothrix species had 2.94% isolation rate each. About 29.6% of the isolated fungi could not be identified. Fungal positive lungs showed acute to sub-acute inflammatory responses with bronchiolar septal, alveolar interstitium and peribronchial inflammatory cell infiltration and thickening of the lung tissues. All the screened fungal isolates were sensitive to voriconazole with the zone of inhibition ranging from 12-27mm. The fungal isolates were all resistant to fluconazole and G. kola extract, except one that showed sensitivity to the extract. Voriconazole showed strong antifungal effect on all the fungi; but the efficacy of the G. kola on the fungal isolates was very low compared to voriconazole.

Multidisciplinary Approach in COVID-19 Pneumonia: A Backward Path by an Expert Team

Background: The novel coronavirus disease 2019 (COVID-19) represents a medical challenge worldwide. COVID-19 pneumonia is an extremely complex disease. The hypothesis of the study was that a multidisciplinary approach involving experienced specialists in diffuse parenchymal lung disease might improve the diagnosis of patients with COVID-19 pneumonia.Methods: Two pulmonologists, two radiologists, and two pathologists reviewed 27 patients who died of severe COVID-19 pneumonia as the main diagnosis made by non-pulmonologists. To evaluate whether the contribution of specialists, individually and/or in combination, might modify the original diagnosis, a three-step virtual process was planned. Pulmonologists, radiologists and pathologists were asked to classify every case into four distinct levels of diagnostic certainty, based on clinical, radiological, and morphological/virologic data obtained from an autoptic lung sample, respectively. The whole lung examination was considered the gold standard for the final diagnosis. The probability of a correct diagnosis was calculated, and the effectiveness of a multidisciplinary diagnosis was obtained by comparing diagnoses made by experienced pulmonologists with those made by non-pulmonologists. Results: COVID-19 pneumonia was excluded in 2 cases (8%) and was a marginal feature in 3 cases (11%). The probability of a correct diagnosis increased strikingly from an undedicated clinician to an expert specialist, becoming progressively more accurate at different steps. Every single specialist made significantly more correct diagnoses than any non-pulmonologist. The highest level of accuracy was achieved by the combination of 3 expert specialists.Conclusions: In summary, the dynamic interaction between expert specialists significantly improves the diagnostic confidence and management of patients with COVID-19 pneumonia.

RNA-seq of a lung sample from a male COVID19 deceased patient (age 74) submitted by Icahn School of Medicine at Mount Sinai does not show the anaerobic/aerobic homeostasis disruption observed in other patient samples - an artifact of post-mortem procedures?

Here, I have analyzed the RNA-seq of a lung sample from a male COVID19 deceased patient (age 74) submitted by Icahn School of Medicine at Mount Sinai (Accid:PRJNA615032). There are two replicates, which I have pooled into a single sample. The analysis (Table 1, SI/bact.txt) does not show the anaero- bic/aerobic homeostasis disruption observed in some patient samples [1–3]. Although not all samples show anaerobic/aerobic homeostasis disruption [4–6], they are possibly patients in the early phase of the disease. The findings in the current deceased patient (viral load is very low, SI/viralreads.fa) however does not align with the hypothesis [7], unless there are artifacts arising from obtaining the lung sample. Since obligate anaerobic bacteria cant tolerate more than 5% oxygen, and atmospheric oxygen is much higher, could these have been killed after atmospheric exposure? Note, anaerobic bacteria are present - its just that they don’t seem to have colonized the lung of the deceased patient.

SARS-Cov2 enables anaerobic bacteria (Prevotella, et al) to colonize the lungs disrupting homeostasis, causing long-drawn chronic symptoms, and acute severe symptoms (ARDS, septic shock, clots, arterial stroke) which finds resonance, with key differences, in the ‘forgotten disease’ Lemierre Syndrome, enabled by Epstein Barr Virus

Metagenomic studies of Covid19 patient sequencing data from different countries (China, Brazil, Peru, Cambodia, USA) shows a pattern that SARS-Cov2 enables anaerobic bacteria (eg Prevotella, Veil- lonella, Capnocytophaga, Fusobacterium, Oribacterium and Bacteroides) colonize the lungs, disrupting the homeostasis found in healthy patients. Long drawn symptoms in Covid19 have caused great con- sternation, and could be explained by persistence of biofilms. Some of these bacteria are implicated in increasing IL-6, cause ground glass opacity in lungs and are associated with cardiac injury - all symp- toms associated with Covid19. Many studies also show several bacterial infection markers - like D-dimer, LDH, C-reactive protein and ferritin - being significantly high, while the viral immune response is at- tenuated (reported by three studies till date). This is also confirmed here in the lung sample from a 74 year old deceased patient, showing high levels of IFITM3, ferritin and S100 calcium binding protein. Anaerobic bacteria causing initial symptoms like persistent fever, chills, pain and later symptoms like ARDS, blood clots, arterial stroke and septic shock finds resonance in a ”forgotten disease” - Lemierre syndrome (LS). While, LS is enabled by Epstein Barr Virus - possibly by ‘a transient depression of T cell immunity’, two recent studies show that IFN-λ might promote bacterial superinfection in Covid19. Also, 16S rRNA bacterial genes and endotoxins (LPS) were discovered in 18/19 severely ill pneumonia patients in one study, suggesting dissemination of endotoxins, and not actual bacteria, might suffice to cause severity. Autopsies also show foci of acute bronchopneumonia. There are key differences with LS - for example origin of LS is the jugular vein while Covid19 starts in the lungs (and this difference should result in Covid19 to be easier to treat). Co-infection of EBV and SARS-Cov2 leads to greater symptoms (fever, higher CRP) in a study of 67 patients. SARS-2003 showed a lot of similar symptoms, which did not get enough media attention. There was a specific warning issued in 2004 for ‘an increased vigilance against stroke and other thrombotic complications among critically-ill SARS patients in future outbreaks’. Enhanced pathogen testing kits, which include RT-PCT for bacterial genes and endotoxin tests, could confirm this disruption in Covid19, and thus anaerobic-specific antibiotics could significantly help in therapy.

Disseminated Mycobacterium kansasii infection in a white-tailed deer and implications for public and livestock health

We document a case of Mycobacterium kansasii, a rare, zoonotic bacterium, in a white-tailed deer (WTD; Odocoileus virginianus) in East Baton Rouge Parish, Louisiana. Grossly, the deer had fibrinous pleuropneumonia with yellow, mineralized nodules scattered throughout the lungs and extending to the pleura. The kidneys were enlarged and had numerous pale foci in the cortex. Microscopically, the pulmonary architecture was replaced by variably sized, multifocal-to-coalescing granulomas with peripheral histiocytes and fewer multinucleate giant cells, and necrotic centers with mineralization and hemorrhage. The latter rarely contained one to a few acid-fast, slender, 7-µm long bacteria, for which beaded morphology was sometimes evident. Similar acid-fast bacteria were also within histiocytes in the kidney. PCR assay of fresh lung sample and subsequent sequencing revealed a non-tuberculosis mycobacterium, M. kansasii. These lesions were similar to those that result from infection with M. bovis in WTD. Both M. bovis and M. kansasii are zoonotic. WTD are a reservoir of M. bovis, which is a major concern in regions in which WTD and cattle can come into close contact.

A Pilot Study of Wet Lung Using Lung Ultrasound Surface Wave Elastography in an Ex Vivo Swine Lung Model

Extravascular lung water (EVLW) is a basic symptom of congestive heart failure and other conditions. Computed tomography (CT) is standard method used to assess EVLW, but it requires ionizing radiation and radiology facilities. Lung ultrasound reverberation artifacts called B-lines have been used to assess EVLW. However, analysis of B-line artifacts depends on expert interpretation and is subjective. Lung ultrasound surface wave elastography (LUSWE) was developed to measure lung surface wave speed. This pilot study aimed at measureing lung surface wave speed due to lung water in an ex vivo swine lung model. The surface wave speeds of a fresh ex vivo swine lung were measured at 100 Hz, 200 Hz, 300 Hz, and 400 Hz. An amount of water was then filled into the lung through its trachea. Ultrasound imaging was used to guide the water filling until significant changes were visible on the imaging. The lung surface wave speeds were measured again. It was found that the lung surface wave speed increases with frequency and decreases with water volume. These findings are confirmed by experimental results on an additional ex vivo swine lung sample.

Differential pulmonary immunopathologic response of domestic sheep (Ovis aries) and bighorn sheep (Ovis canadensis) to Mycoplasma ovipneumoniae infection: a retrospective study

Mycoplasma ovipneumoniae is a respiratory pathogen that can impact domestic sheep (Ovis aries; DS) and bighorn sheep (Ovis canadensis; BHS). Experimental and field data have indicated BHS are more susceptible than DS to developing polymicrobial pneumonia associated with Mycoplasma ovipneumoniae infection. We hypothesized that DS and BHS have a differential immunopathologic pulmonary response to M. ovipneumoniae infection. A retrospective study was performed using formalin-fixed, paraffin-embedded (FFPE) lung tissue from DS and BHS without and with M. ovipneumoniae detected in the lung tissue (n=8 per group). While each M. ovipneumoniae positive lung sample had microscopic changes typical of infection, including hyperplasia of intrapulmonary bronchus-associated lymphoid tissue (BALT) and respiratory epithelium, DS exhibited a more robust and well-organized BALT formation as compared to BHS. Immunohistochemistry was performed with antibodies reactive in FFPE tissues and specific for leukocyte and cytokine markers: T cell marker CD3, B cell markers CD20 and CD79a, macrophage markers CD163 and Iba1, and cytokine IL-17. Digital analysis was used to quantitate chromogen deposition in regions of interest (ROIs), including alveolar and bronchiolar areas, and bronchiolar subregions (epithelium and BALT). Main effects and interaction of species and infection status were analyzed by beta regression and Bonferroni corrections were performed on pairwise comparisons (PBon<0.05 significance). Significant species differences were identified for bronchiolar CD3 (PBon=0.0023) and CD163 (PBon=0.0224), alveolar CD163 (PBon=0.0057), and for IL-17 in each of the ROIs (alveolar: PBon=0.0009; BALT: PBon=0.0083; epithelium: PBon=0.0007). Infected BHS had a higher abundance of bronchiolar CD3 (PBon=0.0005) and CD163 (PBon=0.0162), and alveolar CD163 (PBon=0.0073). While IL-17 significantly increased with infection in BHS BALT (PBon=0.0179) and alveolar (0.0006) ROIs, abundance in DS showed an insignificant decrease in these ROIs and a significant decrease in epithelial abundance (PBon=0.0019). These findings support the hypothesis that DS and BHS have a differential immunopathologic response to M. ovipneumoniae infection.

Cryoprobe biopsy for the diagnosis of acute hypoxemic respiratory failure of undetermined origin

Rationale Acute hypoxemic respiratory failure is a condition that comprises a wide array of entities. Obtaining a histological lung sample might help reach a diagnosis and direct an appropriate treatment in a select group of patients. Objective To describe our experience in the use of cryobiopsy for the diagnosis of acute hypoxemic respiratory failure of undetermined origin. Methods Retrospective analysis of case series of patients with acute hypoxemic respiratory failure who underwent lung cryobiopsy at the Intensive Care Unit of the Hospital Italiano de Buenos Aires, Argentina. Results Cryobiopsy yielded a histological diagnosis in all patients ( n = 10, 100%). This led to either a change in therapy or continuation of a specific treatment in eight of these patients. Cryobiopsy was found to be contributive in all the patients who did not meet Berlin criteria for acute respiratory distress syndrome. No major complications were associated with the procedure. Conclusions Cryobiopsy is a safe procedure with a high diagnostic yield in a selected group of patients.

Molecular identification of Toxoplasma gondii in roadkill wild animals in Yucatan, Mexico

Toxoplasma gondii is an obligate intracellular protozoan parasite, recognized as the etiologic agent of toxoplasmosis, a zoonotic endemic disease in several countries, including Mexico. In the Yucatan State of Mexico, Toxoplasma infection has a high impact in both human and domestic animal health. Wild animals can also host zoonotic pathogens such as Toxoplasma gondii. The presence of Toxoplasma gondii DNA in roadkill wild animals in Yucatan was detected using a nested Polymerase Chain Reaction. Toxoplasma gondii DNA was identified in several organs retrieved from a Yucatan squirrel (Sciurus yucatanensis), a coatimundi (Nasua narica), and a greater grison (Galictis vittata). The amplified fragments of Toxoplasma gondii DNA were purified, sequenced, and certified by BLAST analysis. Our results confirm that Toxoplasma gondii can infect wild mammals from Yucatan, which could act as intermediate hosts and contribute to the transmission of the disease to humans and domestic animals, as well as other wild animal species. We present the first molecular evidence of Toxoplasma gondii in a squirrel and a coatimundi from Yucatan, and quite possibly in a greater grison at a global level.Figure 1. Agarose gel presenting PCR amplicons (560 bp) positive to Toxoplasma gondii. 1) C+: positive control; A: Yucatan squirrel liver sample; B: great grison femoral muscle sample; C: coatimundi kidney sample; C-: negative control. 2) A: Yucatan squirrel brain sample; B: great grison lung sample; C-: negative control.

Utility of Core Needle Biopsies and Transbronchial Biopsies for Diagnosing Nonneoplastic Lung Diseases

Context.— Small lung biopsies (core needle biopsies and transbronchial biopsies) are the most common—and often the first—lung sample obtained when a radiologic abnormality is detected and tissue diagnosis is required. When a neoplastic diagnosis cannot be made but pathologic abnormalities are present, it is useful for pathologists to have a list (“menu”) of specific nonneoplastic diagnoses that can be made in these samples. Objective.— To provide surgical pathologists and pathology trainees with menus of nonneoplastic entities that can be diagnosed in small lung biopsies, and to briefly describe and illustrate some of these entities as they appear in small lung biopsies. Data Sources.— Published literature and the authors' experience with small lung biopsies for diagnosis of nonneoplastic lung diseases. Conclusions.— Although sampling error imposes some limitations, core needle biopsies and transbronchial lung biopsies can contribute to the diagnosis of a variety of nonneoplastic lung diseases and reduce the need for invasive surgical intervention.