Recombinant Paracoccin Reproduces the Biological Properties of the Native Protein and Induces Protective Th1 Immunity against Paracoccidioides brasiliensis Infection

Background

Paracoccin is a dual-function protein of the yeast Paracoccidioides brasiliensis that has lectin properties and N-acetylglucosaminidase activities. Proteomic analysis of a paracoccin preparation from P. brasiliensis revealed that the sequence matched that of the hypothetical protein encoded by PADG-3347 of isolate Pb-18, with a polypeptide sequence similar to the family 18 endochitinases. These endochitinases are multi-functional proteins, with distinct lectin and enzymatic domains.

Methodology/principal findings

The multi-exon assembly and the largest exon of the predicted ORF (PADG-3347), was cloned and expressed in Escherichia coli cells, and the features of the recombinant proteins were compared to those of the native paracoccin. The multi-exon protein was also used for protection assays in a mouse model of paracoccidioidomycosis.

Conclusions/Significance

Our results showed that the recombinant protein reproduced the biological properties described for the native protein—including binding to laminin in a manner that is dependent on carbohydrate recognition—showed N-acetylglucosaminidase activity, and stimulated murine peritoneal macrophages to produce high levels of TNF-α and nitric oxide. Considering the immunomodulatory potential of glycan-binding proteins, we also investigated whether prophylactic administration of recombinant paracoccin affected the course of experimental paracoccidioidomycosis in mice. In comparison to animals injected with vehicle (controls), mice treated with recombinant paracoccin displayed lower pulmonary fungal burdens and reduced pulmonary granulomas. These protective effects were associated with augmented pulmonary levels of IL-12 and IFN-γ. We also observed that injection of paracoccin three days before challenge was the most efficient administration protocol, as the induced Th1 immunity was balanced by high levels of pulmonary IL-10, which may prevent the tissue damage caused by exacerbated inflammation. The results indicated that paracoccin is the protein encoded by PADG-3347, and we propose that this gene and homologous proteins in other P. brasiliensis strains be called paracoccin. We also concluded that recombinant paracoccin confers resistance to murine P. brasiliensis infection by exerting immunomodulatory effects.     

Transmissibility of the Monkeypox Virus Clades via Respiratory Transmission: Investigation Using the Prairie Dog-Monkeypox Virus Challenge System

Monkeypox virus (MPXV) is endemic within Africa where it sporadically is reported to cause outbreaks of human disease. In 2003, an outbreak of human MPXV occurred in the US after the importation of infected African rodents. Since the eradication of smallpox (caused by an orthopoxvirus (OPXV) related to MPXV) and cessation of routine smallpox vaccination (with the live OPXV vaccinia), there is an increasing population of people susceptible to OPXV diseases. Previous studies have shown that the prairie dog MPXV model is a functional animal model for the study of systemic human OPXV illness. Studies with this model have demonstrated that infected animals are able to transmit the virus to naive animals through multiple routes of exposure causing subsequent infection, but were not able to prove that infected animals could transmit the virus exclusively via the respiratory route. Herein we used the model system to evaluate the hypothesis that the Congo Basin clade of MPXV is more easily transmitted, via respiratory route, than the West African clade. Using a small number of test animals, we show that transmission of viruses from each of the MPXV clade was minimal via respiratory transmission. However, transmissibility of the Congo Basin clade was slightly greater than West African MXPV clade (16.7% and 0% respectively). Based on these findings, respiratory transmission appears to be less efficient than those of previous studies assessing contact as a mechanism of transmission within the prairie dog MPXV animal model.     

Effects of helicopter net-gunning on survival of bighorn sheep

Wildlife capture, and the data collection associated with it, has led to major advancements in ecology that are integral to decision making pertaining to wildlife conservation. Capturing wildlife, however, can cause lethal and non-lethal risks to animals. Understanding the factors that contribute to the level of risk involved in wildlife capture is therefore important for the development and implementation of the safest and most effective methodologies. We used data from 736 animal captures of 389 individuals for 2 subspecies of female bighorn sheep (Rocky Mountain bighorn [Ovis canadensis canadensis], Sierra Nevada bighorn sheep [O. c. sierrae]) in Wyoming and California, USA, in 2002–2020 to evaluate the degree and extent of time that capture via helicopter net-gunning affects survival. We compared pre- and post-capture survival during a 10-week window centered on a capture event, and post-capture survival between captured animals and animals that were monitored but not captured during the 10-week window. Additionally, we evaluated the effects of handling techniques (number of times captured, season of capture event, handling time, chase time, and body temp) and biological factors (age and nutritional condition) on probability of capture mortality. Mean daily survival was 0.9992 during a 5-week pre-capture window, dropped to 0.9864 on the day of capture, and rebounded within 3 days of capture to pre-capture levels and that of sheep that were not captured. Overall, direct mortality resulting from capture was 1.36%, with 0.54% mortality occurring within the 3 days following a capture event for an overall 1.90% capture-related mortality. The only handling and biological metrics that influenced the probability of capture mortality were rectal temperature and nutritional condition; high initial rectal temperatures and poor body condition were associated with increased risk of mortality in the days following capture. Overall, helicopter net-gunning imposed low and short-term risk to survival of female bighorn sheep. To reduce bias in survival estimates, we recommend using a 3-day censorship window for post-capture mortalities as opposed to the common practice of a 2–5-week censor window. Helicopter net-gunning, including annual or seasonal recaptures, remains an effective and comparatively safe technique for capture and associated data collection of bighorn sheep.     

Comparative Genotyping of Campylobacter jejuni Strains from Patients with Guillain-Barré Syndrome in Bangladesh

Background

Campylobacter jejuni is a common cause of acute gastroenteritis and is associated with post-infectious neuropathies such as the Guillain-Barré syndrome (GBS) and the Miller Fisher syndrome (MFS). We here present comparative genotyping of 49 C. jejuni strains from Bangladesh that were recovered from patients with enteritis or GBS. All strains were serotyped and analyzed by lipo-oligosaccharide (LOS) genotyping, amplified fragment length polymorphism (AFLP) analysis, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE).

Methodology/Principal Findings

C. jejuni HS:23 was a predominant serotype among GBS patients (50%), and no specific serotype was significantly associated with GBS compared to enteritis. PCR screening showed that 38/49 (78%) of strains could be assigned to LOS classes A, B, C, or E. The class A locus (4/7 vs 3/39; p<0.01) was significantly associated in the GBS-related strains as compared to enteritis strains. All GBS/oculomotor related strains contained the class B locus; which was also detected in 46% of control strains. Overlapping clonal groups were defined by MLST, AFLP and PFGE for strains from patients with gastroenteritis and GBS. MLST defined 22 sequence types (STs) and 7 clonal complexes including 7 STs not previously identified (ST-3742, ST-3741, ST-3743, ST-3748, ST-3968, ST-3969 and ST-3970). C. jejuni HS:23 strains from patients with GBS or enteritis were clonal and all strains belonged to ST-403 complex. Concordance between LOS class B and ST-403 complex was revealed. AFLP defined 25 different types at 90% similarity. The predominant AFLP type AF-20 coincided with the C. jejuni HS:23 and ST-403 complex.

Conclusion/Significance

LOS genotyping, MLST, AFLP and PFGE helped to identify the HS:23 strains from GBS or enteritis patients as clonal. Overall, genotypes exclusive for enteritis or for GBS-related strains were not obtained although LOS class A was significantly associated with GBS strains. Particularly, the presence of a clonal and putative neuropathogenic C. jejuni HS:23 serotype may contribute to the high prevalence of C. jejuni related GBS in Bangladesh.     

Genetic architecture,biochemical underpinnings and ecological impact of floral UV patterning

Floral attraction traits can significantly affect pollinator visitation patterns, but adaptive evolution of these traits may be constrained by correlations with other traits. In some cases, molecular pathways contributing to floral attraction are well characterized, offering the opportunity to explore loci potentially underlying variation among individuals. Here, we quantify the range of variation in floral UV patterning (i.e. UV ‘bulls‐eye nectar guides) among crop and wild accessions of Brassica rapa. We then use experimental crosses to examine the genetic architecture, candidate loci and biochemical underpinnings of this patterning as well as phenotypic manipulations to test the ecological impact. We find qualitative variation in UV patterning between wild (commonly lacking UV patterns) and crop (commonly exhibiting UV patterns) accessions. Similar to the majority of crops, recombinant inbred lines (RILs) derived from an oilseed crop × WI fast‐plant^® cross exhibit UV patterns, the size of which varies extensively among genotypes. In RILs, we further observe strong statistical‐genetic and QTL correlations within petal morphological traits and within measurements of petal UV patterning; however, correlations between morphology and UV patterning are weak or nonsignificant, suggesting that UV patterning is regulated and may evolve independently of overall petal size. HPLC analyses reveal a high concentration of sinapoyl glucose in UV‐absorbing petal regions, which, in concert with physical locations of UV‐trait QTLs, suggest a regulatory and structural gene as candidates underlying observed quantitative variation. Finally, insects prefer flowers with UV bulls‐eye patterns over those that lack patterns, validating the importance of UV patterning in pollen‐limited populations of B. rapa.     

Bartonella spp. Bacteremia in Blood Donors from Campinas,Brazil

Bartonella species are blood-borne, re-emerging organisms, capable of causing prolonged infection with diverse disease manifestations, from asymptomatic bacteremia to chronic debilitating disease and death. This pathogen can survive for over a month in stored blood. However, its prevalence among blood donors is unknown, and screening of blood supplies for this pathogen is not routinely performed. We investigated Bartonella spp. prevalence in 500 blood donors from Campinas, Brazil, based on a cross-sectional design. Blood samples were inoculated into an enrichment liquid growth medium and sub-inoculated onto blood agar. Liquid culture samples and Gram-negative isolates were tested using a genus specific ITS PCR with amplicons sequenced for species identification. Bartonella henselae and Bartonella quintana antibodies were assayed by indirect immunofluorescence. B. henselae was isolated from six donors (1.2%). Sixteen donors (3.2%) were Bartonella-PCR positive after culture in liquid or on solid media, with 15 donors infected with B. henselae and one donor infected with Bartonella clarridgeiae. Antibodies against B. henselae or B. quintana were found in 16% and 32% of 500 blood donors, respectively. Serology was not associated with infection, with only three of 16 Bartonella-infected subjects seropositive for B. henselae or B. quintana. Bartonella DNA was present in the bloodstream of approximately one out of 30 donors from a major blood bank in South America. Negative serology does not rule out Bartonella spp. infection in healthy subjects. Using a combination of liquid and solid cultures, PCR, and DNA sequencing, this study documents for the first time that Bartonella spp. bacteremia occurs in asymptomatic blood donors. Our findings support further evaluation of Bartonella spp. transmission which can occur through blood transfusions.     

Mesoionic compounds with antifungal activity against Fusarium verticillioides

Background

Fungi contaminate the food of humans and animals, are a risk to health, and can cause financial losses. In this work, the antifungal activities of 16 mesoionic compounds (MI 1–16) were evaluated against mycotoxigenic fungi, including Aspergillus spp., Fusarium verticillioides and Penicillium citrinum. Furthermore, the decreased ergosterol in the total lipid content of Fusarium verticillioides was investigated.

Results

F. verticillioides was the most sensitive fungus to the mesoionic compounds. Among the evaluated compounds, MI-11 and MI-16 presented higher antifungal effects against F. verticillioides, with MIC values of 7.8 μg/ml, and MI-2 and MI-3 followed, with MICs of 15.6 μg/ml. The most active compounds were those with heterocyclic ring phenyl groups substituted by electron donor moieties (MI-11 and MI-16). Among some compounds with higher activity (MI-2, MI-11 and MI-16), decreased ergosterol content in the total lipid fraction of F. verticillioides was demonstrated. MI-2 reduced the ergosterol content approximately 40% and 80% at concentrations of 7.8 μg/ml and 15.6 μg/ml, respectively, and MI-11 and MI-16 decreased the content by 30% and 50%, respectively, when at a concentration of 7.8 μg/ml.

Conclusion

These findings indicate that mesoionic compounds have significant antifungal activity against F. verticillioides.     

Pelagic seabird flight patterns are consistent with a reliance on olfactory maps for oceanic navigation

Homing studies have provided tantalizing evidence that the remarkable ability of shearwaters (Procellariiformes) to pinpoint their breeding colony after crossing vast expanses of featureless open ocean can be attributed to their assembling cognitive maps of wind-borne odours but crucially, it has not been tested whether olfactory cues are actually used as a system for navigation. Obtaining statistically important samples of wild birds for use in experimental approaches is, however, impossible because of invasive sensory manipulation. Using an innovative non-invasive approach, we provide strong evidence that shearwaters rely on olfactory cues for oceanic navigation. We tested for compliance with olfactory-cued navigation in the flight patterns of 210 shearwaters of three species (Cory''s shearwaters, Calonectris borealis, North Atlantic Ocean, Scopoli''s shearwaters, C. diomedea Mediterranean Sea, and Cape Verde shearwaters, C. edwardsii, Central Atlantic Ocean) tagged with high-resolution GPS loggers during both incubation and chick rearing. We found that most (69%) birds displayed exponentially truncated scale-free (Lévy-flight like) displacements, which we show are consistent with olfactory-cued navigation in the presence of atmospheric turbulence. Our analysis provides the strongest evidence yet for cognitive odour map navigation in wild birds. Thus, we may reconcile two highly disputed questions in movement ecology, by mechanistically connecting Lévy displacements and olfactory navigation. Our approach can be applied to any species which can be tracked at sufficient spatial resolution, using a GPS logger.     

Perceived Impact of Dental Pain on the Quality of Life of Preschool Children and Their Families

The aim of the present study was to evaluate the perceived impact of dental caries and dental pain on oral health-related quality of life (OHRQoL) among preschool children and their families. A cross-sectional study was conduct with 843 preschool children in Campina Grande, Brazil. Parents/caregivers answered a questionnaire on socio-demographic information, their child’s general/oral health and history of dental pain. The Brazilian version of the Early Childhood Oral Health Impact Scale was administered to determine the perceived impact of caries and dental pain on OHRQoL. The children underwent an oral examination. Logistic regression for complex sample was used to determine associations between the dependent and independent variables (OR: Odds ratio, α = 5%). The independents variables that had a p-value <0.20 in the bivariate analysis were selected for the multivariate model. The prevalence of dental caries and dental pain was 66.3% and 9.4%, respectively. Order of birth of the child, being the middle child (OR: 10.107, 95%CI: 2.008-50.869) and youngest child (OR: 3.276, 95%CI: 1.048-10.284) and dental pain (OR: 84.477, 95%CI: 33.076-215.759) were significant predictors of the perceived impact on OHRQOL for children. Poor perception of oral health was significant predictor of the perceived impact on OHRQOL for family (OR=7.397, 95%CI: 2.190-24.987). Dental caries was not associated with a perceived impact on the ORHQoL of either the children or their families. However, order of child birth and dental pain were indicators of impact of OHRQoL on preschool children and poor perception of oral health was indicators of impact on families.     

Terpenes of Podocarpus lambertius

Sitosterol and the following terpenic compounds have been isolated from the bark of Podocarpus lambertius: 3β-hydroxytotarol, 4β-carboxynortotarol, and macrophyllic and lambertic acids. The leaves yielded sitosterol, stigmastan-3β,5α-diol-6-one, isopimaric acid, phyllocladene, isophyllocladene, 8,9-abieten-15-ol and 17-isophyllocladenol.