Pathogens — Open Access Journal

The rapidly evolving coronavirus disease 2019 (COVID-19, caused by severe acute respiratory syndrome coronavirus 2- SARS-CoV-2), has greatly burdened the global healthcare system and led it into crisis in several countries. Lack of targeted therapeutics led to the idea of repurposing broad-spectrum drugs for viral intervention. In vitro analyses of hydroxychloroquine (HCQ)’s anecdotal benefits prompted its widespread clinical repurposing globally. Reports of emerging cardiovascular complications due to its clinical prescription are revealing the crucial role of angiotensin-converting enzyme 2 (ACE2), which serves as a target receptor for SARS-CoV-2. In the present settings, a clear understanding of these targets, their functional aspects and physiological impact on cardiovascular function are critical. In an up-to-date format, we shed light on HCQ’s anecdotal function in stalling SARS-CoV-2 replication and immunomodulatory activities. While starting with the crucial role of ACE2, we here discuss the impact of HCQ on systemic cardiovascular function, its associated risks, and the scope of HCQ-based regimes in current clinical settings. Citing the extent of HCQ efficacy, the key considerations and recommendations for the use of HCQ in clinics are further discussed. Taken together, this review provides crucial insights into the role of ACE2 in SARS-CoV-2-led cardiovascular activity, and concurrently assesses the efficacy of HCQ in contemporary clinical settings. Full article

Mycoplasma bovis is an important bovine pathogen causing pneumonia, mastitis, and arthritis and is responsible for major economic losses worldwide. In the absence of an efficient vaccine, control of M. bovis infections mainly relies on antimicrobial treatments, but resistance is reported in an increasing number of countries. To address the situation in Spain, M. bovis was searched in 436 samples collected from beef and dairy cattle (2016–2019) and 28% were positive. Single-locus typing using polC sequences further revealed that two subtypes ST2 and ST3, circulate in Spain both in beef and dairy cattle, regardless of the regions or the clinical signs. Monitoring of ST2 and ST3 isolates in a minimum inhibitory concentration (MIC) to a panel of antimicrobials revealed one major difference when using fluoroquinolones (FQL): ST2 is more susceptible than ST3. Accordingly, whole-genome sequencing (WGS) further identified mutations in the gyrA and parC regions, encoding quinolone resistance-determining regions (QRDR) only in ST3 isolates. This situation shows the capacity of ST3 to accumulate mutations in QRDR and might reflect the selective pressure imposed by the extensive use of these antimicrobials. MIC values and detection of mutations by WGS also showed that most Spanish isolates are resistant to macrolides, lincosamides, and tetracyclines. Valnemulin was the only one effective, at least in vitro, against both STs. Full article

Bacteria colonizing the mouth induce an adaptive immune response with the systemic and local presence of species or strain-specific immunoglobulins. Few studies have addressed global antibody patterns for oral bacteria or potential population time trends. We assessed these aspects in relation to a panel of oral bacteria. Using multiplex immunoblotting, IgG levels for 26 oral bacterial species (54 strains) were determined in 888 plasma samples from 30-year-old early pregnant women (n = 516) and 50-year-old men and women (n = 372) collected between 1976 and 2018. Inter-species correlations were found and age-dependent profiles and levels of immune responses to oral bacteria confirmed. We found temporal trends in the global and single-species antibody responses, but this was age-specific with both inclining and declining shifts. Prominent shifts in the younger group increased IgG towards health-associated Streptococcus salivarius and Streptococcus sanguinis, and in the older group towards disease-associated Aggregatibacter actinomycetemcomitans, Filifactor alocis, and Streptococcus mutans, among others. We concluded that temporal shifts occurred from 1976 to 2018, which may reflect improved oral health (more remaining teeth) and altered lifestyle habits, but this needs to be evaluated in observational studies considering more aspects. Full article

The human T-cell lymphotropic virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia/lymphoma (ATLL), a neoplasm of CD4+CD25+ T cells that occurs in 2–5% of infected individuals after decades of asymptomatic latent infection. Multiple HTLV-1-encoded regulatory proteins, including Tax and HTLV-1 basic leucine zipper factor (HBZ), play key roles in viral persistence and latency. The HTLV-1 Tax oncoprotein interacts with a plethora of host cellular proteins to regulate viral gene expression and also promote the aberrant activation of signaling pathways such as NF-κB to drive clonal proliferation and survival of T cells bearing the HTLV-1 provirus. Tax undergoes various post-translational modifications such as phosphorylation and ubiquitination that regulate its function and subcellular localization. Tax shuttles in different subcellular compartments for the activation of anti-apoptotic genes and deregulates the cell cycle with the induction of DNA damage for the accumulation of genomic instability that can result in cellular immortalization and malignant transformation. However, Tax is highly immunogenic and therefore HTLV-1 has evolved numerous strategies to tightly regulate Tax expression while maintaining the pool of anti-apoptotic genes through HBZ. In this review, we summarize the key findings on the oncogenic mechanisms used by Tax that set the stage for the development of ATLL, and the strategies used by HTLV-1 to tightly regulate Tax expression for immune evasion and viral persistence. Full article

The pathogen Phytophthora infestans is responsible for catastrophic crop damage on a global scale which totals billions of euros annually. The discovery of new inhibitors of this organism is of paramount agricultural importance and of critical relevance to food security. Current strategies for crop treatment are inadequate with the emergence of resistant strains and problematic toxicity. Natural products such as cinnamaldehyde have been reported to have fungicidal properties and are the seed for many new discovery research programmes. We report a probe of the cinnamaldehyde framework to investigate the aldehyde subunit and its role in a subset of aromatic aldehydes in order to identify new lead compounds to act against P. infestans. An ellipticine derivative which incorporates an aldehyde (9-formyl-6-methyl ellipticine, 34) has been identified with exceptional activity versus P. infestans with limited toxicity and potential for use as a fungicide. Full article

The hard tick Dermacentor reticulatus transmits Babesia canis, the causative agent of canine babesiosis. Both the occurrence and local distribution of D. reticulatus as well as infection rates of questing ticks with B. canis are thus far poorly known in Bavaria, Germany. The objectives of this study were to conduct (1) a georeferenced field study on the occurrence of D. reticulatus with digital habitat characterization and (2) a PCR analysis of D. reticulatus collected in Bavaria for infection with B. canis. Dermacentor reticulatus were collected by flagging at 60 sites specifically selected according to habitat conditions and screened individually for Babesia DNA. A digital habitat characterization for D. reticulatus was performed according to results of the field analysis including the parameters land use, proximity to water, “potential natural vegetation”, red deer corridors and climate data. Altogether, 339 D. reticulatus ticks (214 females and 125 males) were collected between 2010 and 2013 at 12 out of 60 sampling sites. All 12 sites were characterized by high humidity with marshy areas. Babesia canis DNA was detected in 1 out of 301 (0.3%) questing D. reticulatus in Bavaria. The digital habitat characterization revealed 15 forest areas in Bavaria with similar ecological characteristics as the sites positive for D. reticulatus. Full article

Puumala orthohantavirus (PUUV) causes nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome (HFRS) commonly diagnosed in Europe. The majority of HFRS cases in the European part of Russia are diagnosed in the Volga Federal District, which includes the Republic of Tatarstan (RT). The current study aims to analyze the genetic variability of PUUV in Pre-Kama region of the RT bounded by the Volga, Kama, and Vyatka rivers. In 2017, bank voles were caught in seven isolated forest traps in the Pre-Kama region and for the 26 PUUV-positive samples, the partial small (S), medium (M), and large (L) genome segment sequences were obtained and analyzed. It was determined that all identified PUUV strains belong to the Russian (RUS) genetic lineage; however, the genetic distance between strains is not directly correlated with the geographical distance between bank vole populations. One of the identified strains has S and L segments produced from one parental strain, while the M segment was supplied by another, suggesting that this strain could be the reassortant. We suggest that the revealed pattern of the PUUV strains distribution could be the result of a series of successive multidirectional migratory flows of the bank voles to the Pre-Kama region in the postglacial period. Full article

The sequential pathology of a genotype XIII Bangladeshi strain of Newcastle disease virus (NDV) was studied in 5-weeks old chickens. Layer chickens of ISA Brown breed were inoculated through the intranasal and intraocular routes with the BD-C161/2010 strain of NDV and examined at different times post-infection (pi). NDV-infected chickens showed depression at 3 days pi (dpi) followed by dropped wings, paralysis and death starting at 4 dpi. Lungs of infected chickens showed hemorrhagic lesions starting at 24 hours pi (hpi) that was followed by pallor and slight contraction by 2 to 3 dpi and subsequently developed into severe hemorrhagic pneumonia with mononuclear cell infiltration. Hemorrhagic and necrotizing lesions were found in different visceral organs including proventriculus, intestine, gut-associated lymphoid tissues, liver and kidneys starting at 3 dpi that progressed rapidly. Severe lymphoid depletion was observed in the thymus, spleen and bursa of Fabricius starting at 1–3 dpi followed by hemorrhages, necrosis, inflammation and atrophy at 4–5 dpi. In the brain, mild neuronal lesions such as focal to diffuse encephalitis with encephalomalacia was observed at 2–3 dpi and moderate and diffuse meningoencephalitis with encephalomalacia at advanced stages. In conclusion, the BD-C161/2010 strain of NDV produced lesions typical of velogenic viscerotropic pathotype of NDV. Full article

In the absence of therapeutic interventions, and a possible vaccine candidate, the spread of COVID-19 disease and associated fatalities are on the rise. The high mutation frequency in the genomic material of these viruses supports their ability to adapt to new environments, resulting in an efficient alteration in tissue tropism and host range. Therefore, the coronavirus’ health threats could be relevant for the long-term. The epidemiological data indicate that age, sex, and cardio-metabolic disease have a significant impact on the spread and severity of COVID-19. In this review, we highlight recent updates on the pathogenesis of SARS-CoV-2 among men and women, including children. We also discuss the role of the cellular receptors and coreceptors used by the virus to enter host cells on differential infection among men, women, and cardio-metabolic patients. Full article

Italian beef production is mainly based on a feedlot system where calves are housed with mixed aged cattle often in conditions favourable to bovine respiratory disease (BRD). In Veneto, an indoor system is also used for imported bulls around 300–350 kg. Mycoplasmas, in particular Mycoplasma bovis and Mycoplasma dispar, contribute to BRD in young calves, but their role in the disease in older cattle has not been investigated. In this study, ten heads of cattle were selected from each of the 24 groups kept in 13 different farms. Bulls were sampled by nasal swabbing at 0, 15, and 60 days after arrival for Mycoplasma isolation. Identification was carried out by 16S-rDNA PCR followed by denaturing gradient gel electrophoresis. M. bovis, M. dispar, and M. bovirhinis were identified, and prevalence was analysed by mixed-effects logistic regression models. This showed that most bulls arrived free of M. bovis, but within two weeks, approximately 40% became infected, decreasing to 13% by the last sampling. In contrast, the prevalence of M. dispar was not dependent on time or seasonality, while M. bovirhinis only showed a seasonality-dependent trend. The Italian fattening system creates an ideal environment for infection with M. bovis, probably originating from previously stabled animals. Full article

Previous studies have revealed that the electrically charged disinfectant CAC-717 has strong virucidal and bactericidal effects but is safe for humans and animals. In this study, CAC-717 was further evaluated for its potential effects as a disinfectant against scrapie prions. Western blotting showed that CAC-717 reduced the amount of the abnormal isoform of prion protein (PrP^Sc) in prion-infected cell (ScN2a) lysates. Furthermore, the reduction of prion transmissibility was confirmed by a mouse bioassay, in which mice injected with scrapie prions pre-treated with CAC-717 survived longer than those injected with untreated scrapie prions. Lastly, to evaluate the seeding activity of ScN2a cell lysates treated with CAC-717, quantitative protein misfolding cyclic amplification (PMCA) was performed directly on ScN2a cell lysates treated with CAC-717, which showed that the median dose of PMCA (PMCA₅₀) dropped from log9.95 to log5.20 after CAC-717 treatment, indicating more than a 4 log reduction. This suggests that the seeding activity of PrP^Sc is decreased by CAC-717. Collectively, these results suggest that CAC-717 has anti-prion activity, reducing both PrP^Sc conversion activity and prion transmissibility; thus, CAC-717 will be useful as a novel disinfectant in prion diseases. Full article

Feline infectious peritonitis (FIP) is a common infectious cause of death in cats, with heritable host factors associated with altered risk of disease. To assess the role of feline interferon-gamma gene (fIFNG) variants in this risk, the allele frequencies of two single nucleotide polymorphisms (SNPs) (g.401 and g.408) were determined for non-pedigree cats either with confirmed FIP (n = 59) or from the general population (cats enrolled in a large lifetime longitudinal study; n = 264). DNA was extracted from buccal swabs or tissue samples. A pyrosequencing assay to characterize the fIFNG SNPs was designed, optimized and subsequently performed on all samples. Genotype and allele frequency were calculated for each population. Characterization of the target SNPs was possible for 56 of the cats with FIP and 263 of the cats from the general population. The SNPs were in complete linkage disequilibrium with each other. There was an association between FIP status and genotype (χ²; p = 0.028), with a reduced risk of developing FIP (χ²; p = 0.0077) associated with the genotype TT at both positions. These results indicate that, although fIFNG variants may be associated with altered risk of disease, the prevalence of individual variants within both populations limits application of their characterization to breeding purposes. Full article

Species designations within Acanthamoeba are problematic because of pleomorphic morphology. Molecular approaches, including DNA sequencing, hinted at a resolution that has yet to be fully achieved. Alternative approaches were required. In 1996, the Byers/Fuerst lab introduced the concept of sequence types. Differences between isolates of Acanthamoeba could be quantitatively assessed by comparing sequences of the nuclear 18S rRNA gene, ultimately producing 22 sequence types, designated T1 through T22. The concept of sequence types helps our understanding of Acanthamoeba evolution. Nevertheless, substantial variation in the 18S rRNA gene differentiates many isolates within each sequence type. Because the majority of isolates with sequences in the international DNA databases have been studied for only a small segment of the gene, designated ASA.S1, genetic variation within this hypervariable region of the 18S rRNA gene has been scrutinized. In 2002, we first categorized variation in this region in a sample of T3 and T4 isolates from Hong Kong, observing ten “alleles” within type T4 and five “alleles” within T3. Subsequently, confusion occurred when different labs applied redundant numerical labels to identify different alleles. A more unified approach was required. We have tabulated alleles occurring in the sequences submitted to the international DNA databases, and determined their frequencies. Over 150 alleles have occurred more than once within 3500+ isolates of sequence type T4. Results from smaller samples of other sequence types (T3, T5, T11 and T15, and supergroup T2/6) have also been obtained. Our results provide new insights into the evolutionary history of Acanthamoeba, further illuminating the degree of genetic separation between significant taxonomic units within the genus, perhaps eventually elucidating what constitutes a species of Acanthamoeba. Full article

Porcine circovirus 3 (PCV-3) has been suggested as a putative causal agent of swine reproductive disease. A number of different studies have pointed out this association, but there is still a lack of information regarding the normal rates of PCV-3 infection in farms with normal reproductive parameters. The objective of the present study was to assess the frequency of PCV-3 detection in primiparous and multiparous sows and in tissues from their respective fetuses from farms with average reproductive parameters. Sera from 57 primiparous and 64 multiparous sows from 3 different farms were collected at two time points. Brain and lung tissues from 49 mummies and 206 stillborn were collected at farrowing. Samples were tested by PCR, and when positive, quantified by quantitative PCR. Thirty-nine complete genomes were obtained and phylogenetically analyzed. All sera from multiparous sows were negative, while 19/57 (33.3%) primiparous sows were PCV-3 PCR positive. From the 255 tested fetuses, 86 (33.7%) had at least one tissue positive to PCV-3. The frequency of detection in fetuses from primiparous sows (73/91, 80.2%) was significantly higher than those from multiparous ones (13/164, 7.9%). It can be concluded that PCV-3 is able to cause intrauterine infections in absence of overt reproductive disorders. Full article

Campylobacter jejuni is a major foodborne pathogen and common cause of bacterial enteritis worldwide. A total of 622 C. jejuni isolates recovered from food animals and retail meats in the United States through the National Antimicrobial Resistance Monitoring System between 2013 and 2017 were sequenced using an Illumina MiSeq. Sequences were combined with WGS data of 222 human isolates downloaded from NCBI and analyzed by core genome multilocus sequence typing (cgMLST) and traditional MLST. cgMLST allelic difference (AD) thresholds of 0, 5, 10, 25, 50, 100 and 200 identified 828, 734, 652, 543, 422, 298 and 197 cgMLST types among the 844 isolates, respectively, and traditional MLST identified 174 ST. The cgMLST scheme allowing an AD of 200 (cgMLST₂₀₀) revealed strong correlation with MLST. cgMLST₂₀₀ showed 40.5% retail chicken isolates, 56.5% swine, 77.4% dairy cattle and 78.9% beef cattle isolates shared cgMLST sequence type with human isolates. All ST-8 had the same cgMLST₂₀₀ type (cgMLST₂₀₀-12) and 74.3% of ST-8 and 75% cgMLST₂₀₀-12 were confirmed as sheep abortion virulence clones by PorA analysis. Twenty-nine acquired resistance genes, including 21 alleles of bla_OXA, tetO, aph(3′)-IIIa, ant(6)-Ia, aadE, aad9, aph(2′)-Ig, aph(2′)-Ih, sat4 plus mutations in gyrA, 23SrRNA and L22 were identified. Resistance genotypes were strongly linked with cgMLST₂₀₀ type for certain groups including 12/12 cgMLST₂₀₀-510 with the A103V substitution in L22 and 10/11 cgMLST₂₀₀-608 with the T86I GyrA substitution associated with macrolide and quinolone resistance, respectively. In summary, the cgMLST₂₀₀ threshold scheme combined with resistance genotype information could provide an excellent subtyping scheme for source attribution of human C. jejuni infections. Full article

Maize/soybean relay strip intercropping has been widely practiced in Southwest China due to its high productivity and effective application of agricultural resources; however, several seedborne diseases such as seedling blight, pod and seed decay are frequently observed causing severe yield loss and low seed quality. So far, the population and pathogenicity of the seedborne fungi associated with intercropped soybean remain unexplored. In this study, seeds of 12 soybean cultivars screened for intercropping were collected from three growing regions in Sichuan Province of Southwest China, and the seedborne fungi were isolated from the surface-sterilized seeds. Based on sequence analysis of ribosomal DNA internal transcribed spacer (rDNA ITS), 148 isolates were identified into 13 fungal genera, among which Fusarium covered 55.0% as the biggest population followed by Colletotrichum. Furthermore, Fusarium isolates were classified into five distinct species comprising F. fujikuroi, F. proliferatum, F. verticillioides, F. asiaticum and F. incarnatum through sequence analysis of translation elongation factor 1 alpha (EF-1α) and DNA-directed RNA ploymerase II second largest subunit (RPB2). Among them, F. fujikuroi accounted for 51.22% (42/82) and was isolated from 91.7% (11/12) soybean varieties. Pathogenicity assay showed that five Fusarium species were able to infect the seeds of soybean cultivar “Nandou12” and caused water-soaked or rot symptoms, while F. fujikuroi and F. asiaticum had much higher aggressiveness than other species with significant reductions of seed fresh weight and germination percentage. Accordingly, this study indicates that Fusarium species are the dominant seedborne fungi in the intercropped soybean in Sichuan, China, and this provides some useful references for the effective management of seedborne fungal diseases as well as soybean resistance breeding in maize/soybean relay strip intercropping. Full article

(1) Background: Studying neutrophils in vitro is difficult since these cells are terminally differentiated and are easily activated during isolation. At the same time, most of the available model cell lines are associated with certain limitations, such as functional deficiency or a lack of expression of surface markers characteristic of neutrophils. P. gingivalis is a periodontopathogen that causes dysbiosis in subgingival bacterial biofilm. This triggers the accumulation of functional neutrophils in the periodontium. However, until now, the specific effects of P. gingivalis-derived lipopolysaccharide on neutrophil functions have not been analyzed. (2) Methods: The impact of two variants of commercially available P. gingivalis endotoxin on neutrophil functions was tested using the HoxB8 in vitro system that is well suited to analyze neutrophil response to different stimuli in a controlled manner. (3) Results: The Standard P. gingivalis lipopolysaccharide (LPS), known to activate cells through Toll-like receptor 2 (TLR2)- and Toll-like receptor 4 (TLR4)-dependent pathways, prolonged neutrophil survival and exhibited pro-inflammatory effects. In contrast, Ultrapure LPS, binding exclusively to TLR4, neither protected neutrophils from apoptosis, nor induced an inflammatory response. (4) Conclusion: Two variants of P. gingivalis-derived LPS elicited effects on neutrophils and, based on the obtained results, we concluded that the engagement of both TLR2 and TLR4 is required for the manipulation of survival and the stimulation of immune responses of HoxB8 neutrophils. Full article

COVID-19 is the first known pandemic caused by a coronavirus, SARS-CoV-2, which is the third virus in the family Coronaviridae to cause fatal infections in humans after SARS-CoV and MERS-CoV. Animals are involved in the COVID-19 pandemic. This review summarizes the role of animals as reservoirs, natural hosts and experimental models. SARS-CoV-2 originated from animal reservoir, most likely bats and/or pangolins. Anthroponotic transmission has been reported in cats, dogs, tigers, lions and minks. As of now, there is no a strong evidence for natural animal-to-human transmission or sustained animal-to-animal transmission of SARS-CoV-2. Experimental infections conducted by several research groups have shown that monkeys, hamsters, ferrets, cats, tree shrews, transgenic mice and fruit bats were permissive, while dogs, pigs and poultry were resistant. There is an urgent need to understand the zoonotic potential of different viruses in animals, particularly in bats, before they transmit to humans. Vaccines or antivirals against SARS-CoV-2 should be evaluated not only for humans, but also for the protection of companion animals (particularly cats) and susceptible zoo and farm animals. Full article

Listeria monocytogenes is a ubiquitous opportunistic pathogen responsible for the well-known listeriosis disease. This bacterium has become a common contaminant of food, threatening the food processing industry. Once consumed, the pathogen is capable of traversing epithelial barriers, cellular invasion, and intracellular replication through the modulation of virulence factors such as internalins and haemolysins. Mobile genetic elements (plasmids and transposons) and other sophisticated mechanisms are thought to contribute to the increasing antimicrobial resistance of L. monocytogenes. The environmental persistence of the pathogen is aided by its ability to withstand environmental stresses such as acidity, cold stress, osmotic stress, and oxidative stress. This review seeks to give an insight into L. monocytogenes biology, with emphasis on its virulence factors, antimicrobial resistance, and adaptations to environmental stresses. Full article