Microorganisms, Volume 9, Issue 5 (May 2021) – 228 articles

The influenza A virus (IAV) PA-X protein is a virulence factor that selectively degrades host mRNAs leading to protein shutoff. This function modulates host inflammation, antiviral responses, cell apoptosis, and pathogenesis. In this work we describe a novel approach based on the use of bacteria and plasmid encoding of the PA-X gene under the control of the bacteriophage T7 promoter to identify amino acid residues important for A/Brevig Mission/1/1918 H1N1 PA-X’s shutoff activity. Using this system, we have identified PA-X mutants encoding single or double amino acid changes, which diminish its host shutoff activity, as well as its ability to counteract interferon responses upon viral infection. This novel bacteria-based approach could be used for the identification of viral proteins that inhibit host gene expression as well as the amino acid residues responsible for inhibition of host gene expression. Full article

This study aims to determine differences in the on- and off-tumor microbiota between patients with right- and left-sided colorectal cancer. Microbiome profiling of tumor and tumor-adjacent biopsies from patients with right-sided (n = 17) and left-sided (n = 7) colorectal adenocarcinoma was performed using 16S ribosomal RNA sequencing. Off-tumor alpha and beta diversity were significantly different between right- and left-sided colorectal cancer patients. However, no differences in on-tumor diversity were observed between tumor locations. Comparing the off-tumor microbiota showed the right colon to be enriched with species of the Lachnoclostridium, Selenomonas, and Ruminococcus genera. Whereas the left colon is enriched with Epsilonbacteraeota phylum, Campylobacteria class, and Pasteurellales and Campylobacterales orders, in contrast, the on-tumor microbiota showed relatively fewer differences in bacterial taxonomy between tumor sites, with left tumors being enriched with Methylophilaceae and Vadin BE97 families and Alloprevotella, Intestinibacter, Romboutsia, and Ruminococcus 2 genera. Patients with left-sided colorectal cancer had large taxonomic differences between their paired on- and off-tumor microbiota, while patients with right-sided colorectal cancer showed relatively fewer taxonomic differences. Collectively, this suggests that the right and left colon show distinctive bacterial populations; however, the presence of a colonic tumor leads to a more consistent microbiota between locations. Full article

Agrobacterium tumefaciens is a pathogen of various plants which transfers its own DNA (T-DNA) to the host plants. It is used for producing genetically modified plants with this ability. To control T-DNA transfer to the right place, toxin-antitoxin (TA) systems of A. tumefaciens were used to control the target site of transfer without any unintentional targeting. Here, we describe a toxin-antitoxin system, Atu0939 (mazE-at) and Atu0940 (mazF-at), in the chromosome of Agrobacterium tumefaciens. The toxin in the TA system has 33.3% identity and 45.5% similarity with MazF in Escherichia coli. The expression of MazF-at caused cell growth inhibition, while cells with MazF-at co-expressed with MazE-at grew normally. In vivo and in vitro assays revealed that MazF-at inhibited protein synthesis by decreasing the cellular mRNA stability. Moreover, the catalytic residue of MazF-at was determined to be the 24th glutamic acid using site-directed mutagenesis. From the results, we concluded that MazF-at is a type II toxin-antitoxin system and a ribosome-independent endoribonuclease. Here, we characterized a TA system in A. tumefaciens whose understanding might help to find its physiological function and to develop further applications. Full article

The objective of the study was to compare three quantification methods to a “garbage can audit” (reference method, REF) for monitoring antimicrobial usage (AMU) from products other than medicated feed over one year in 101 Québec dairy farms. Data were collected from veterinary invoices (VET method), from the “Amélioration de la Santé Animale au Québec” provincial program (GOV method), and from farm treatment records (FARM method). The AMU rate was reported in a number of Canadian Defined Course Doses for cattle (DCDbovCA) per 100 cow-years. Electronic veterinary sales data were obtained for all farms for VET and GOV methods. For the FARM method, a herd management software was used by 68% of producers whereas farm treatment records were handwritten for the others; records could not be retrieved in 4% of farms. Overall, agreement was almost perfect between REF and VET methods (concordance correlation coefficient (CCC) = 0.83), but moderate between REF and GOV (CCC = 0.44), and between REF and FARM (CCC = 0.51). Only a fair or slight agreement was obtained between any alternative method of quantification and REF for oral and intrauterine routes. The billing software used by most of Québec’s dairy veterinary practitioners seems promising in terms of surveillance and benchmarking of AMU in the province. Full article

Due to the increasing reports of carbapenemase-producing Enterobacteriaceae (CPE) from livestock in recent years, the European Reference Laboratory for Antimicrobial Resistances (EURL-AR) provided a protocol for their recovery from caecum and meat samples. This procedure exhibited limitations for the detection of CPE with low carbapenem MIC values. Therefore, it was modified by a second, selective enrichment in lysogeny broth with cefotaxime (CTX 1 mg/L) and with meropenem (MEM 0.125 mg/L) at 37 °C under microaerophilic conditions. By Real-time PCR, these enrichments are pre-screened for the most common carbapenemase genes. Another adaptation was the use of in-house prepared MacConkey agar with MEM and MEM+CTX instead of commercial selective agar. According to the EURL-method, we achieved 100% sensitivity and specificity using the in-house media instead of commercial agar, which decreased the sensitivity to ~75%. Comparing the method with and without the second enrichment, no substantial influence on sensitivity and specificity was detected. Nevertheless, this enrichment has simplified the CPE-isolation regarding the accompanying microbiota and the separation of putative colonies. In conclusion, the sensitivity of the method can be increased with slight modifications. Full article

Kuroshio Current intrusion (KCI) has significant impacts on the oceanographic conditions and ecological processes of the Pacific-Asian marginal seas. Little is known to which extent and how, specifically, the microzooplankton community can be influenced through the intrusion. Here, we focused on ciliates that often dominated the microzooplankton community and investigated their communities using high-throughput sequencing of 18S rRNA gene transcripts in the northern South China Sea (NSCS), where the Kuroshio Current (KC) intrudes frequently. We first applied an isopycnal mixing model to assess the fractional contribution of the KC to the NSCS. The ciliate community presented a provincial distribution pattern corresponding to more and less Kuroshio-influenced stations. Structural equation modeling revealed a significant impact of the KCI on the community, while environmental variables had a marginal impact. KCI-sensitive OTUs were taxonomically diverse but mainly belonged to classes Spirotrichea and Phyllopharyngea, suggesting the existence of core ciliates responding to the KCI. KCI-sensitive OTUs were grouped into two network modules that showed contrasting abundance behavior with the KC fraction gradient, reflecting differential niches (i.e., winner and loser) in the ciliate community during the Kuroshio intrusion scenarios. Our study showed that the Kuroshio intrusion, rather than environmental control, was particularly detrimental to the oligotrophic microzooplankton community. Full article

Melting polar and alpine ice surfaces frequently exhibit blooms of dark pigmented algae. These microbial extremophiles significantly reduce the surface albedo of glaciers, thus accelerating melt rates. However, the ecology, physiology and taxonomy of cryoflora are not yet fully understood. Here, a Swiss and an Austrian glacier dominated either by filamentous Ancylonema nordenskioeldii or unicellular Mesotaenium berggrenii var. alaskanum, were sampled. Molecular analysis showed that both species are closely related, sharing identical chloroplast morphologies (parietal-lobed for Ancylonema vs. axial plate-like for Mesotaenium sensu stricto), thus the unicellular species was renamed Ancylonema alaskana. Moreover, an ecophysiological comparison of the two species was performed: pulse–amplitude modulated (PAM) fluorometry confirmed that they have a high tolerance to elevated solar irradiation, the physiological light preferences reflected the conditions in the original habitat; nonetheless, A. nordenskioeldii was adapted to higher irradiances while the photosystems of A. alaskana were able to use efficiently low irradiances. Additionally, the main vacuolar polyphenol, which effectively shields the photosystems, was identical in both species. Also, about half of the cellular fatty acids were polyunsaturated, and the lipidome profiles dominated by triacylglycerols were very similar. The results indicate that A. alaskana is physiologically very similar and closely related but genetically distinct to A. nordenskioeldii. Full article

Since biofilm formation by microfoulers significantly contributes to the fouling process, it is important to evaluate the performance of marine surfaces to prevent biofilm formation, as well as understand their interactions with microfoulers and how these affect biofilm development and structure. In this study, the long-term performance of five surface materials—glass, perspex, polystyrene, epoxy-coated glass, and a silicone hydrogel coating—in inhibiting biofilm formation by cyanobacteria was evaluated. For this purpose, cyanobacterial biofilms were developed under controlled hydrodynamic conditions typically found in marine environments, and the biofilm cell number, wet weight, chlorophyll a content, and biofilm thickness and structure were assessed after 49 days. In order to obtain more insight into the effect of surface properties on biofilm formation, they were characterized concerning their hydrophobicity and roughness. Results demonstrated that silicone hydrogel surfaces were effective in inhibiting cyanobacterial biofilm formation. In fact, biofilms formed on these surfaces showed a lower number of biofilm cells, chlorophyll a content, biofilm thickness, and percentage and size of biofilm empty spaces compared to remaining surfaces. Additionally, our results demonstrated that the surface properties, together with the features of the fouling microorganisms, have a considerable impact on marine biofouling potential. Full article

Leishmaniasis are neglected diseases caused by several species of Leishmania that affect humans and many domestic and wild animals with a worldwide distribution. The objectives of this review are to identify wild animals naturally infected with zoonotic Leishmania species as well as the organs infected, methods employed for detection and percentage of infection. A literature search starting from 1990 was performed following the PRISMA methodology and 161 reports were included. One hundred and eighty-nine species from ten orders (i.e., Carnivora, Chiroptera, Cingulata, Didelphimorphia, Diprotodontia, Lagomorpha, Eulipotyphla, Pilosa, Primates and Rodentia) were reported to be infected, and a few animals were classified only at the genus level. An exhaustive list of species; diagnostic techniques, including PCR targets; infected organs; number of animals explored and percentage of positives are presented. L. infantum infection was described in 98 wild species and L. (Viania) spp. in 52 wild animals, while L. mexicana, L. amazonensis, L. major and L. tropica were described in fewer than 32 animals each. During the last decade, intense research revealed new hosts within Chiroptera and Lagomorpha. Carnivores and rodents were the most relevant hosts for L. infantum and L. (Viannia) spp., with some species showing lesions, although in most of the studies clinical signs were not reported. Full article

The bat tick Cariosvespertilionis has been reported from Sweden to occasionally feed on humans resulting in disease symptoms. The aim of this study was to investigate C. vespertilionis as a potential vector and reservoir of Borrelia species. In 2015 and 2018 in south-central Sweden, C. vespertilionis ticks were collected from a wooden bat box harboring Soprano pipistrelle bats, Pipistrellus pygmaeus. In addition, one C. vespertilionis tick found inside a house in southern Sweden in 2019 was collected. Ticks were screened for Borrelia spp. using a genus-specific quantitative PCR assay. The Borrelia species of the positive specimens were determined by conventional PCR followed by DNA sequencing and phylogenetic analyses. A total of 24% (22 of 92) of the analyzed C. vespertilionis ticks were Borrelia-positive. Phylogenetic analyses indicate that the bacteria belong to the relapsing fever group of borreliae; some of them appear to be identical with Borrelia sp. CPB1, a spirochete only found twice before—in the United Kingdom and in France. Our results also indicate a temporal and spatial distribution of this Borrelia species. Since C. vespertilionis occasionally bites humans, and since it exhibits a high prevalence of Borrelia bacteria, it is possible that it presents a risk of human disease. Further studies are needed to characterize Borrelia sp. CPB1 to determine if it is human-pathogenic and to determine if C. vespertilionis is a vector and/or reservoir of this agent. Full article

Ensiling has been widely applied to cope with agricultural solid waste to achieve organic waste valorization and relieve environmental pressure and feedstuff shortage. In this study, co-ensiling of cabbage leaf byproduct and rice straw was performed with inoculation of Lactiplantibacillusplantarum (LP) to investigate the effects of inoculation on ensiling performance and microflora profiles. Compared to the control, LP inoculation preserved more dry matter (DM) content (283.4 versus 270.9 g·kg⁻¹ fresh matter (FM) on day 30), increased lactic acid (LA) content (52.1 versus 35.8 g·kg⁻¹ dry matter on day 15), decreased pH (3.55 versus 3.79 on day 15), and caused accumulation of acetic acid (AA), butyric acid (BA), and ammonia. The investigation showed that LP inoculation modified microflora composition, especially resisting potential pathogens and enriching more lactic acid bacteria (LAB) (p < 0.05). Moreover, Lactiplantibacillus and Lacticaseibacillus were identified as the keystone taxa that influenced physicochemical properties and interactions in microflora. They were also the main functional species that directly restrained undesirable microorganisms (p < 0.05), rather than indirectly working via metabolite inhibition and substrate competition (p > 0.05). The results of this present study improve the understanding of the underlying effect of LP inoculation on improving silage quality and facilitate the bio-transformation of cabbage byproduct and rice straw as animal feed. Full article

Obesity is a complex syndrome and is recognized as the ultimate pathway of many chronic diseases. Studies using Akkermansia muciniphila supplementation strategy have proved to be effective for the prevention and treatment of obesity and other metabolic disorders. Although there are studies that support the protective effect of this strategy, the effects on the prevention of obesity on humans are not clear yet and need more investigation. The aim of this study is to investigate the effect of A. muciniphila administration on modulating obesity. This systematic review was generated from articles published within the last 10 years. All articles were in English and included animal subjects. The review relied on the search engines Google Scholar, Pub Med, Web of Science and Medline using the following keywords: A. muciniphila, next-generation probiotic, new-generation probiotic, obesity, fat mass, body fat and lipid profile. The search has revealed 804 articles with relevant key words. After the exclusion of irrelevant articles, 10 studies were selected based on the criteria. These studies were randomized controlled trials that have shown that A. muciniphila modulates obesity by regulating metabolism and energy hemostasis and improving insulin sensitivity and glucose hemostasis. In addition, studies showed this microorganism enhances low grade inflammation by different mechanisms. Full article

Neospora caninum and Toxoplasma gondii affect both humans and animals worldwide. To investigate their seroprevalence and differences in seropositivity between pigs vaccinated and unvaccinated against porcine circovirus 2 (PCV2), as well as differences in muscle enzyme activity between seropositive and seronegative pigs, blood samples were collected from 380 sows. Antibodies against T. gondii and N. caninum were detected by an indirect immunofluorescence antibody (IFA) assay, while the activities of creatine kinase (CK) and aspartate aminotransferase (AST) were biochemically assessed. Out of the 364 sows finally included in the study, 4.4%, 3.5%, and 0.5% were seropositive to T. gondii, N. caninum, or both. A significantly higher percentage of seropositivity against T. gondii and/or N. caninum in PCV2 unvaccinated pigs compared with vaccinated pigs was observed. Increased serum activities of CK and AST were detected in 71.43% and 100% of only against T. gondii (T+) and 63.64% and 90.91% of only against N. caninum (N+) seropositive sows, respectively, and were significantly higher compared to seronegative animals. T. gondii and N. caninum seropositivity, especially in presumed immunocompromised pigs, and the evidence of muscle damage highlight their importance as a zoonotic pathogen and animal model of human infection, respectively. Full article

The isolation and molecular and chemo-taxonomic identification of seventeen halophilic archaea from the Santa Bárbara saltern, Sonora, México, were performed. Eight strains were selected based on pigmentation. Molecular identification revealed that the strains belonged to the Haloarcula, Halolamina and Halorubrum genera. Neutral lipids (quinones) were identified in all strains. Glycolipid S-DGD was found only in Halolamina sp. strain M3; polar phospholipids 2,3-O-phytanyl-sn-glycerol-1-phosphoryl-3-sn-glycerol (PG), 2,3-di-O-phytanyl-sn-glycero-1-phospho-3′-sn-glycerol-1′-methyl phosphate (PGP-Me) and sodium salt 1-(3-sn-phosphatidyl)-rac-glycerol were found in all the strains; and one unidentified glyco-phospholipid in strains M1, M3 and M4. Strains M1, M3 and M5 were selected for further studies based on carotenoid production. The effect of glucose and succinic and glutamic acid on carotenoid production was assessed. In particular, carotenoid production and growth significantly improved in the presence of glucose in strains Haloarcula sp. M1 and Halorubrum sp. M5 but not in Halolamina sp. M3. Glutamic and succinic acid had no effect on carotenoid production, and even was negative for Halorubrum sp. M5. Growth was increased by glutamic and succinic acid on Haloarcula sp. M1 but not in the other strains. This work describes for first time the presence of halophilic archaea in the Santa Bárbara saltern and highlights the differences in the effect of carbon sources on the growth and carotenoid production of haloarchaea. Full article

Streptococcus suis is an important zoonotic pathogen causing severe infections in swine and humans. Induction of the Vibrio parahaemolyticus YoeB toxin in Escherichia coli resulted in cell death, leading to the speculation that YoeB_Vp can be a counterselectable marker. Herein, the counterselection potential of YoeB_Vp was assessed in S. suis. The yoeB_Vp gene was placed under the copper-induced promoter PcopA. The PcopA-yoeB_Vp construct was cloned into the S. suis-E. coli shuttle vector pSET2 and introduced into S. suis to assess the effect of YoeB_Vp expression on S. suis growth. Reverse transcription quantitative PCR showed that copper induced yoeB_Vp expression. Growth curve analyses and spot dilution assays showed that YoeB_Vp expression inhibited S. suis growth both in liquid media and on agar plates, revealing that YoeB_Vp has the potential to be a counterselectable marker for S. suis. A SCIY cassette comprising the spectinomycin-resistance gene and copper-induced yoeB_Vp was constructed. Using the SCIY cassette and peptide-induced competence, a novel two-step markerless gene deletion method was established for S. suis. Moreover, using the ΔperR mutant generated by this method, we demonstrated that PmtA, a ferrous iron and cobalt efflux pump in S. suis, was negatively regulated by the PerR regulator. Full article

A variety of antiviral treatments for COVID-19 have been investigated, involving many repurposed drugs. Currently, the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp, encoded by nsp12-nsp7-nsp8) has been targeted by numerous inhibitors, e.g., remdesivir, the only provisionally approved treatment to-date, although the clinical impact of these interventions remains inconclusive. However, the potential emergence of antiviral resistance poses a threat to the efficacy of any successful therapies on a wide scale. Here, we propose a framework to monitor the emergence of antiviral resistance, and as a proof of concept, we address the interaction between RdRp and remdesivir. We show that SARS-CoV-2 RdRp is under purifying selection, that potential escape mutations are rare in circulating lineages, and that those mutations, where present, do not destabilise RdRp. In more than 56,000 viral genomes from 105 countries from the first pandemic wave, we found negative selective pressure affecting nsp12 (Tajima’s D = −2.62), with potential antiviral escape mutations in only 0.3% of sequenced genomes. Potential escape mutations included known key residues, such as Nsp12:Val473 and Nsp12:Arg555. Of the potential escape mutations involved globally, in silico structural models found that they were unlikely to be associated with loss of stability in RdRp. No potential escape mutation was found in a local cohort of remdesivir treated patients. Collectively, these findings indicate that RdRp is a suitable drug target, and that remdesivir does not seem to exert high selective pressure. We anticipate our framework to be the starting point of a larger effort for a global monitoring of drug resistance throughout the COVID-19 pandemic. Full article

The recovery of yeasts and lactic acid bacteria (LAB) involved in sourdough fermentation is the first step in the selection of starters with suitable technological aptitude and capable of producing desired aromas and/or aromatic precursors. In this work, two sourdoughs samples (MA and MB) and the derived doughs (samples A and B) were collected from a bakery during artisanal Panettone manufacture. Yeasts and bacteria were isolated at different fermentation steps on selective agar media. A total of 77 isolates were obtained and characterized. Representative strains of yeasts and LAB were identified by sequencing the D1/D2 domain of the 26S rRNA and the 16S rRNA genes, respectively. Moreover, the volatile organic compounds (VOCs) produced in the collected samples were detected and correlated to the species found in the same samples. The results highlighted the occurrence of Kazachstania humilis in both samples A and B, while Saccharomyces cerevisiae strains were detected only in samples B. Among LAB, Fructilactobacillus sanfranciscensis was the main species detected in both sourdoughs. Furthermore, strains belonging to the species Lactiplantibacillus plantarum, Furfurilactobacillus rossiae, Lactobacillus parabuchneri, Leuconostoc citreum, and Leuconostoc mesenteroides were assessed in the dough samples. Full article

The Asia-Pacific hosts a large diversity of canine vector-borne pathogens (VBPs) with some of the most common and most pathogenic, generating significant mortality as well as a spectrum of health impacts on local dog populations. The VBPs Anaplasma platys, Babesia gibsoni, Babesia vogeli, Ehrlichia canis, Hepatozoon canis and haemotropic Mycoplasma spp. are all endemic throughout the region, with many exhibiting shifting geographical distributions that warrant urgent attention. Moreover, many of these species cause similar clinical signs when parasitising canine hosts, whilst knowledge of the exact pathogen is critical to ensure treatment is effective. This is complicated by frequent coinfection that can exacerbate pathology. Here, we describe the development, optimisation and validation of two novel quadruplex Taq-Man based real-time PCRs (qPCRs) for the specific and sensitive detection of the aforementioned VBPs. To ensure accurate evaluation of diagnostic performance, results of our qPCRs were evaluated on field samples from Thai dogs and compared with both conventional PCR (cPCR) results and next-generation sequencing (NGS) metabarcoding. Our qPCRs were found to be more sensitive at detecting canine VBP than cPCR and generated results similar to those achieved by NGS. These qPCRs will provide a valuable high-throughput diagnostic tool available to epidemiologists, researchers and clinicians for the diagnosis of key canine VBPs in the Asia-Pacific and further afield. Full article

In Alberta’s Athabasca oil sands region (AOSR), over 1.25 billion m³ of tailings waste from the bitumen extraction process are stored in tailings ponds. Fugitive emissions associated with residual hydrocarbons in tailings ponds pose an environmental concern and include greenhouse gases (GHGs), reduced sulphur compounds (RSCs), and volatile organic compounds (VOCs). Froth treatment tailings (FTT) are a specific type of tailings waste stream from the bitumen froth treatment process that contains bioavailable diluent: either naphtha or paraffins. Tailings ponds that receive FTT are associated with the highest levels of biogenic gas production, as diverse microbial communities biodegrade the residual diluent. In this review, current literature regarding the composition, chemical analysis, and microbial degradation of FTT and its constituents is presented in order to provide a more complete understanding of the complex chemistry and biological processes related to fugitive emissions from tailings ponds receiving FTT. Characterizing the composition and biodegradation of FTT is important from an environmental perspective to better predict emissions from tailings ponds and guide tailings pond management decisions. Full article

Parageobacillus thermoglucosidasius is known to catalyse the biological water gas shift (WGS) reaction, a pathway that serves as a source of alternative energy and carbon to a wide variety of bacteria. Despite increasing interest in this bacterium due to its ability to produce biological hydrogen through carbon monoxide (CO) oxidation, there are no data on the effect of toxic CO gas on its physiology. Due to its general requirement of O₂, the organism is often grown aerobically to generate biomass. Here, we show that carbon monoxide (CO) induces metabolic changes linked to distortion of redox balance, evidenced by increased accumulation of organic acids such as acetate and lactate. This suggests that P. thermoglucosidasius survives by expressing several alternative pathways, including conversion of pyruvate to lactate, which balances reducing equivalents (oxidation of NADH to NAD⁺), and acetyl-CoA to acetate, which directly generates energy, while CO is binding terminal oxidases. The data also revealed clearly that P. thermoglucosidasius gained energy and grew during the WGS reaction. Combined, the data provide critical information essential for further development of the biotechnological potential of P. thermoglucosidasius. Full article

The heat shock protein 90 (Hsp90) is thought to be an excellent drug target against parasitic diseases. The leishmanicidal effect of an Hsp90 inhibitor, 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), was previously demonstrated in both in vitro and in vivo models of cutaneous leishmaniasis. Parasite death was shown to occur in association with severe ultrastructural alterations in Leishmania, suggestive of autophagic activation. We hypothesized that 17-AAG treatment results in the abnormal activation of the autophagic pathway, leading to parasite death. To elucidate this process, experiments were performed using transgenic parasites with GFP-ATG8-labelled autophagosomes. Mutant parasites treated with 17-AAG exhibited autophagosomes that did not entrap cargo, such as glycosomes, or fuse with lysosomes. ATG5-knockout (Δatg5) parasites, which are incapable of forming autophagosomes, demonstrated lower sensitivity to 17-AAG-induced cell death when compared to wild-type (WT) Leishmania, further supporting the role of autophagy in 17-AAG-induced cell death. In addition, Hsp90 inhibition resulted in greater accumulation of ubiquitylated proteins in both WT- and Δatg5-treated parasites compared to controls, in the absence of proteasome overload. In conjunction with previously described ultrastructural alterations, herein we present evidence that treatment with 17-AAG causes abnormal activation of the autophagic pathway, resulting in the formation of immature autophagosomes and, consequently, incidental parasite death. Full article

L3 larvae of anisakid nematodes are an important problem for the fisheries industry and pose a potential risk for human health by acting as infectious agents causing allergies and as potential vectors of pathogens and microrganisms. In spite of the close bacteria–nematode relationship very little is known of the anisakids microbiota. Fresh fish could be contaminated by bacteria vectored in the cuticle or in the intestine of anisakids when the L3 larvae migrate through the muscles. As a consequence, the bacterial inoculum will be spread, with potential effects on the quality of the fish, and possible clinical effects cannot be discarded. A total of 2,689,113 16S rRNA gene sequences from a total of 113 L3 individuals obtained from fish captured along the FAO 27 fishing area were studied. Bacteria were taxonomically characterized through 1803 representative operational taxonomic units (OTUs) sequences. Fourteen phyla, 31 classes, 52 orders, 129 families and 187 genera were unambiguously identified. We have found as part of microbiome an average of 123 OTUs per L3 individual. Diversity indices (Shannon and Simpson) indicate an extraordinary diversity of bacteria at an OTU level. There are clusters of anisakids individuals (samples) defined by the associated bacteria which, however, are not significantly related to fish hosts or anisakid taxa. This suggests that association or relationship among bacteria in anisakids, exists without the influence of fishes or nematodes. The lack of relationships with hosts of anisakids taxa has to be expressed by the association among bacterial OTUs or other taxonomical levels which range from OTUs to the phylum level. There are significant biological structural associations of microbiota in anisakid nematodes which manifest in clusters of bacteria ranging from phylum to genus level, which could also be an indicator of fish contamination or the geographic zone of fish capture. Actinobacteria, Aquificae, Firmicutes, and Proteobacteria are the phyla whose abundance value discriminate for defining such structures. Full article

RNA viruses play a significant role in the current high losses of pollinators. Although many studies have focused on the epidemiology of western honey bee (Apis mellifera) viruses at the colony level, the dynamics of virus infection within colonies remains poorly explored. In this study, the two main variants of the ubiquitous honey bee virus DWV as well as three major honey bee viruses (SBV, ABPV and BQCV) were analyzed from Varroa-destructor-parasitized pupae. More precisely, RT-qPCR was used to quantify and compare virus genome copies across honey bee pupae at the individual and subfamily levels (i.e., patrilines, sharing the same mother queen but with different drones as fathers). Additionally, virus genome copies were compared in cells parasitized by reproducing and non-reproducing mite foundresses to assess the role of this vector. Only DWV was detected in the samples, and the two variants of this virus significantly differed when comparing the sampling period, colonies and patrilines. Moreover, DWV-A and DWV-B exhibited different infection patterns, reflecting contrasting dynamics. Altogether, these results provide new insight into honey bee diseases and stress the need for more studies about the mechanisms of intra-colonial disease variation in social insects. Full article

Recently, we reported that nicotine plays a role in the failure of the macrophage in the clearance of Mycobacterium avium subspecies paratuberculosis (MAP) during infection in Crohn’s disease smokers. We also demonstrated that nicotine enhances macrophages cellular survival during MAP infection. Blocking α7 nicotinic acetylcholine receptor (α7nAChR) with the pharmacological antagonist—mecamylamine—subverted the anti-inflammatory effect of nicotine in macrophages. Yet, it is still unknown how α7nAChR is involved in the modulation of the macrophage response during MAP infection. Here, we studied the mechanistic role of nicotine-α7nAChR interaction in modulating NF-ĸB survival pathway, autophagy, and effect on cathelicidin production in MAP-infected macrophages using THP-1 cell lines. Our results showed that nicotine upregulated α7nAChR expression by 5-folds during MAP infection compared to controls. Bcl-2 expression was also significantly increased after nicotine exposure. Moreover, Nicotine inhibited autophagosome formation whereas infection with MAP in absence of nicotine has significantly increased LC-3b in macrophages. Nicotine also further upregulated NF-ĸB subunits expression including Rel-B and p100, and increased nuclear translocation of p52 protein. We also discovered that cathelicidin production was significantly suppressed in MAP-infected macrophages, treatment with nicotine showed no effect. Overall, the study provides new insight toward understanding the cellular role of nicotine through α7nAChR/NF-ĸB p100/p52 signaling pathway in inducing anti-apoptosis and macrophage survival during MAP infection in Crohn’s disease smokers. Full article

Three out of one hundred eighty putative LAB isolates from Korean traditional fermented soybean paste were identified to be unique and bacteriocinogenic strains. Based on phenotypic and 16S rRNA sequencing analysis, selected strains were identified as Enterococcus faecium ST651ea, E. faecium ST7119ea and E. faecium ST7319ea. The bacteriocinogenic properties of the studied strains were evaluated against Listeria monocytogenes ATCC15313, Listeria innocua ATCC33090 and vancomycin-resistant E. faecium VRE19 of clinical origin. The strains E. faecium ST651ea, ST7119ea and ST7319ea expressed bacteriocins with an activity of 12,800 AU/mL, 25,600 AU/mL and 25,600 AU/mL, respectively, recorded against L. monocytogenes ATCC15131. According to the PCR-based screening of bacteriocin-related genes, which was further confirmed through amplicon sequencing, showed that strain E. faecium ST651ea carries entB and entP genes, whereas both E. faecium ST7119ea and ST7319ea strains harbor entA and entB genes. The molecular size of expressed bacteriocins was estimated by tricine-SDS-PAGE showing an approximative protein size of 4.5 kDa. The assessment of the spectrum of activity of bacteriocins ST651ea, ST7119ea and ST7319ea showed strong activity against most of clinical VRE isolates, majority of other Enterococcus spp. and Listeria spp. Bacteriocins ST651ea, ST7119ea and ST7319ea were partially purified by combination of 60% ammonium sulfate precipitation and hydrophobic chromatography on the SepPakC₁₈ column. Challenge test with semi-purified (60% 2-propanol fraction) bacteriocins resulted in a significant reduction of viable cells for all test organisms. Thus, indicating that all the bacteriocins evaluated can be used as potential biocontrol in food and feed industries as well as an alternative treatment for VRE-related infections in both veterinary and clinical settings. Full article

Host-parasite interaction can result in a strong alteration of the host-associated microbiota. This dysbiosis can affect the fitness of the host; can modify pathogen interaction and the outcome of diseases. Biomphalaria glabrata is the snail intermediate host of the trematode Schistosoma mansoni, the agent of human schistosomiasis, causing hundreds of thousands of deaths every year. Here, we present the first study of the snail bacterial microbiota in response to Schistosoma infection. We examined the interplay between B. glabrata, S. mansoni and host microbiota. Snails were infected and the microbiota composition was analysed by 16S rDNA amplicon sequencing approach. We demonstrated that the microbial composition of water did not affect the microbiota composition. Then, we characterised the Biomphalaria bacterial microbiota at the individual scale in both naive and infected snails. Sympatric and allopatric strains of parasites were used for infections and re-infections to analyse the modification or dysbiosis of snail microbiota in different host-parasite co-evolutionary contexts. Concomitantly, using RNAseq, we investigated the link between bacterial microbiota dysbiosis and snail anti-microbial peptide immune response. This work paves the way for a better understanding of snail/schistosome interaction and should have critical consequences in terms of snail control strategies for fighting schistosomiasis disease in the field. Full article

Serratia marcescens is an opportunistic nosocomial pathogen and causes wound and burn infections. It shows high resistance to antibiotics and its pathogenicity is mediated by an arsenal of virulence factors. Another therapeutic option to such infections is targeting quorum sensing (QS), which controls the expression of different S. marcescens virulence factors. Prevention of QS can deprive S. marcescens from its bacterial virulence without applying stress on the bacterial growth and facilitates the eradication of the bacteria by immunity. The objective of the current study is to explore the antimicrobial and antivirulence activities of xylitol against S. marcescens. Xylitol could inhibit the growth of S. marcescens. Sub-inhibitory concentrations of xylitol could inhibit biofilm formation, reduce prodigiosin production, and completely block protease activity. Moreover, xylitol decreased swimming motility, swarming motility and increased the sensitivity to hydrogen peroxide. The expression of rsmA, pigP, flhC, flhD fimA, fimC, shlA bsmB, and rssB genes that regulate virulence factor production was significantly downregulated by xylitol. In silico study showed that xylitol could bind with the SmaR receptor by hydrophobic interaction and hydrogen bonding, and interfere with the binding of the natural ligand with SmaR receptor. An in vivo mice survival test confirmed the ability of xylitol to protect mice against the virulence of S. marcescens. In conclusion, xylitol is a growth and virulence inhibitor in S. marcescens and can be employed for the treatment of S. marcescens wound and burn infections. Full article

Polyomaviruses infect many species, including humans. So far, 15 polyomaviruses have been described in humans, but it remains to be established whether all of these are genuine human polyomaviruses. The most recent polyomavirus to be detected in a person is Quebec polyomavirus (QPyV), which was identified in a metagenomic analysis of a stool sample from an 85-year-old hospitalized man. We used PCR to investigate the presence of QPyV DNA in urine samples from systemic lupus erythematosus (SLE) patients (67 patients; 135 samples), multiple sclerosis patients (n = 35), HIV-positive patients (n = 66) and pregnant women (n = 65). Moreover, cerebrospinal fluid from patients with suspected neurological diseases (n = 63), nasopharyngeal aspirates from patients (n = 80) with respiratory symptoms and plasma samples from HIV-positive patients (n = 65) were examined. QPyV DNA was found in urine from 11 (16.4%), 10 (15.4%) and 5 (14.3%) SLE patients, pregnant women, and multiple sclerosis patients, respectively. No QPyV DNA could be detected in the other samples. Alignment with the only available QPyV sequence in the GenBank revealed amino acid substitutions in the HI-loop of capsid protein VP1 in 6/28 of the isolates. Our results show that QPyV viruria can occur, but whether it may cause clinical symptoms in the patients remains to be determined. Full article

Adding massive amounts of lactic starters to raw milk to manage the sanitary risk in the cheese-making process could be detrimental to microbial diversity. Adjusting the amount of the lactic starter used could be a key to manage these adverse impacts. In uncooked pressed cheeses, we investigated the impacts of varying the doses of a lactic starter (the recommended one, 1×, a 0.1× lower and a 2× higher) on acidification, growth of Staphylococcus aureus SA15 and Shiga-toxin-producing Escherichia coli (STEC) O26:H11 F43368, as well as on the bacterial community patterns. We observed a delayed acidification and an increase in the levels of pathogens with the 0.1× dose. This dose was associated with increased richness and evenness of cheese bacterial community and higher relative abundance of potential opportunistic bacteria or desirable species involved in cheese production. No effect of the increased lactic starter dose was observed. Given that sanitary criteria were paramount to our study, the increase in the pathogen levels observed at the 0.1× dose justified proscribing such a reduction in the tested cheese-making process. Despite this, the effects of adjusting the lactic starter dose on the balance of microbial populations of potential interest for cheese production deserve an in-depth evaluation. Full article