A smelly business: microbiology of Adélie penguin guano (Point Thomas rookery, Antarctica)

Adélie penguins (Pygoscelis adeliae) are the most numerous flightless bird group breeding in coastal areas of Maritime and Continental Antarctica. Their activity leaves a mark on the land in the form of large guano deposits. This guano is an important nutrient source for terrestrial habitats of ice-free Antarctic areas, most notably by being the source of ammonia vapors which feed the surrounding grass, lichen and algae communities. Although investigated by researchers, the fate of the guano-associated microbial community and its role in decomposition processes remain vague. Therefore, by employing several direct community assessment methods combined with a broad culture-based approach we provide data on bacterial numbers, their activity and taxonomic affiliation in recently deposited and decayed Adélie penguin guano sampled at the Point Thomas rookery in Maritime Antarctica (King George Island). Our research indicates that recently deposited guano harbored mostly bacteria of penguin gut origin, presumably inactive in cold rookery settings. This material was rich in mesophilic enzymes active also at low temperatures, likely mediating early stage decomposition. Fresh guano colonization by environmental bacteria was minor, accomplished mostly by ammonia scavenging Jeotgalibaca sp. cells. Decayed guano contained 10-fold higher bacterial numbers with cold-active enzymes dominating the samples. Guano was colonized by uric-acid degrading and lipolytic Psychrobacter spp. and proteolytic Chryseobacterium sp. among others. Several spore-forming bacteria of penguin gut origin persisted in highly decomposed material, most notably uric-acid fermenting members of the Gottschalkiaceae family

Evaluation of the Escherichia coli HK82 and BS87 strains as tools for AlkB studies

Within a decade the family of AlkB dioxygenases has been extensively studied as a one-protein DNA/RNArepair system in Escherichia coli but also as a group of proteins of much wider functions in eukaryotes.Two strains, HK82 and BS87, are the most commonly used E. coli strains for the alkB gene mutations. Theaim of this study was to assess the usefulness of these alkB mutants in different aspects of research onAlkB dioxygenases that function not only in alkylated DNA repair but also in other metabolic processes incells. Using of HK82 and BS87 strains, we found the following differences among these alkB−derivatives:(i) HK82 has shown more than 10-fold higher MMS-induced mutagenesis in comparison to BS87; (ii)different specificity of Arg+revertants; (iii) increased induction of SOS and Ada responses in HK82; (iv)the genome of HK82, in comparison to AB1157 and BS87, contains additional mutations: nalA, sbcC, andnuoC. We hypothesize that in HK82 these mutations, together with the non-functional AlkB protein, mayresult in much higher contents of ssDNA, thus higher in comparison to BS87 MMS-induced mutagenesis.In the light of our findings, we strongly recommend using BS87 strain in AlkB research as HK82, bearingseveral additional mutations in its genome, is not an exact derivative of the AB1157 strain, and showsadditional features that may disturb proper interpretation of obtained results

The Ability of Lytic Staphylococcal Podovirus vB_SauP_phiAGO1.3 to Coexist in Equilibrium With Its Host Facilitates the Selection of Host Mutants of Attenuated Virulence but Does Not Preclude the Phage Antistaphylococcal Activity in a Nematode Infection Model

Phage vB_SauP_phiAGO1.3 (phiAGO1.3) is a polyvalent Staphylococcus lytic podovirus with a 17.6-kb genome (Gozdek et al., 2018). It can infect most of the Staphylococcus aureus human isolates of dominant clonal complexes. We show that a major factor contributing to the wide host range of phiAGO1.3 is a lack or sparcity of target sites for certain restriction-modification systems of types I and II in its genome. Phage phiAGO1.3 requires for adsorption β-O-GlcNAcylated cell wall teichoic acid, which is also essential for the expression of methicillin resistance. Under certain conditions an exposure of S. aureus to phiAGO1.3 can lead to the establishment of a mixed population in which the bacteria and phages remain in equilibrium over multiple generations. This is reminiscent of the so called phage carrier state enabling the co-existence of phage-resistant and phage-sensitive cells supporting a continuous growth of the bacterial and phage populations. The stable co-existence of bacteria and phage favors the emergence of phage-resistant variants of the bacterium. All phiAGO1.3-resistant cells isolated from the phage-carrier-state cultures contained a mutation inactivating the two-component regulatory system ArlRS, essential for efficient expression of numerous S. aureus virulence-associated traits. Moreover, the mutants were unaffected in their susceptibility to infection with an unrelated, polyvalent S. aureus phage of the genus Kayvirus. The ability of phiAGO1.3 to establish phage-carrier-state cultures did not preclude its antistaphylococcal activity in vivo in an S. aureus nematode infection model. Taken together our results suggest that phiAGO1.3 could be suitable for the therapeutic application in humans and animals, alone or in cocktails with Kayvirus phages. It might be especially useful in the treatment of infections with the majority of methicillin-resistant S. aureus strains

Highly Resistant Serotype 19A Streptococcus pneumoniae of the GPSC1/CC320 Clone from Invasive Infections in Poland Prior to Antipneumococcal Vaccination of Children

Abstract Introduction The introduction of pneumococcal conjugate vaccines (PCV) into the national immunization programs (NIPs) has significantly reduced the number of pneumococcal infections. However, infections caused by isolates of non-vaccine serotypes (NVT) started spreading shortly thereafter and strains of NVT 19A have become the main cause of invasive pneumococcal disease burden worldwide. The aim of the study was to characterize serotype 19A invasive pneumococci of GPSC1/CC320 circulating in Poland before the introduction of PCV into the Polish NIP in 2017 and to compare them to isolates from other countries where PCVs were implemented much earlier than in Poland. Methods All the GPSC1/CC320 isolates were analyzed by serotyping, susceptibility testing, and whole genome sequencing followed by analyses of resistome, virulome, and core genome multilocus sequence typing (cgMLST), including comparative analysis with isolates with publicly accessible genomic sequences (PubMLST). Results During continuous surveillance the NRCBM collected 4237 invasive Streptococcus pneumoniae isolates between 1997 and 2016, including 200 isolates (4.7%) of serotype 19A. The most prevalent among 19A pneumococci were highly resistant representatives of Global Pneumococcal Sequence Cluster 1/Clonal Complex 320, GPSC1/CC320 (n = 97, 48.5%). Isolates of GPSC1/CC320 belonged to three sequence types (STs): ST320 (75.2%) ST4768 (23.7%), and ST15047 (1.0%), which all represented the 19A-III cps subtype and had complete loci for both PI-1 and PI-2 pili types. On the basis of the cgMLST analysis the majority of Polish GPSC1/CC320 isolates formed a group clearly distinct from pneumococci of this clone observed in other countries. Conclusion Before introduction of PCV in the Polish NIP we noticed an unexpected increase of serotype 19A in invasive pneumococcal infections, with the most common being representatives of highly drug-resistant GPSC1/CC320 clone, rarely identified in Europe both before and even after PCV introduction

Sensitivity of <i>P. putida</i> AlkA-, AlkB-, and Ada-deficient mutants to alkylating chemicals MMS and MNNG.

<p>The serial dilution drop test of <i>P</i>. <i>putida</i> mutants: 1 – wild-type, 2 – alkB‾, 3 – alkA‾, 4 – ada‾, 5 – alkB‾ada‾, 6 – alkA‾ada‾, 7 – alkA‾alkB‾, 8 – alkA‾alkB‾ada‾ on the medium containing the indicated concentrations of MMS (A) or MNNG (B). The vertical arrow shows the direction of dilution: from 10^-1 to 10^-8.</p

Complementation assay for EcAlkB protein.

<p>Frequency of MMS-induced Arg⁺ revertants in the <i>E</i>. <i>coli</i> alkB‾ strains harboring pVB1x plasmids expressing EcAlkB or the PpAlkB homolog (‘“empty”’ vector served as control) (A). Survival of MMS (B, D) or CAA (C) treated M13 (B, C) or MS2 (D) phages in the same strains.</p

The consensus sequences responsible for Ada protein specific interaction (A) located in promoter sequences (up to -100 nucleotide residues relative to ATG start codons) of <i>E. coli ada</i>, <i>alkA</i>, and <i>aidB</i>.

<p>Additionally, the predicted consensuses in promoter regions of <i>E. coli </i><i>alkB</i> and <i>P. putida </i><i>alkA</i>, <i>gcdH</i>, <i>alkB</i>, and <i>ada</i> genes are shown (green bars). Also, the putative -35 and -10 boxes responsible for RNA polymerase interaction are marked in pink (retrieved from RegulonDB, <a href="http://regulondb.ccg.unam.mx" target="_blank">http://regulondb.ccg.unam.mx</a>) or grey (SoftBerry BPROM prediction tool, <a href="http://linux1.softberry.com" target="_blank"><u>http://linux1.softberry.com</u></a>). The phylogenetic tree (B) shows relative conservation between depicted consensus sequences based on UPGMA Geneious Tree Builder, A box – light blue, B box – brown, numbers indicate substitutions per site (for details see Materials and Methods).</p

The pH dependence of repair rates of the studied adducts by PpAlkB protein.

<p>Samples were assayed for extent of repair at marked pH values at 30°C. The adducts and the conditions applied for their repair were as follows: 3meC (pKa 8.7; Ueda et al. (1963) J Amer Chem 85: 4024), 100 µM Fe(II) 5 pmol PpAlkB, 5 min reaction (A) HPC (pKa 8.6; Borys-Brzywczy et al. (2005) Acta Biochim Pol 52: 149), 100 µM Fe(II) 20 pmol PpAlkB, 15 min reaction (B); HEC (pKa 5.8; Krzyzosiak et al. (1979) Polish J Chem 53: 243), 1 mM Fe(II) 20 pmol PpAlkB, 15 min reaction (C) ;εA (pKa 3.9; Secrist et al. (1972) Biochemistry 11: 3499), 3mM Fe(II) 40 pmol PpAlkB, 15 min reaction (D); and εC (pKa 3.7; Krzyzosiak et al. (1979) Polish J Chem 53: 243), 50 µM Fe(II) 80 pmol PpAlkB, 15 min reaction (E).</p

Dependence of 3meC repair by PpAlkB on the reaction temperature.

<p>Samples were assayed for extent of repair at marked temperature values.</p

Survival of <i>P. putida</i> (A, C) and <i>E. coli</i> (B, D) mutants defective in <i>alkA</i>, <i>alkB</i>, and <i>ada</i> genes treated with 20 mM MMS (A, B) or 5 µg/ml MNNG (C, D) for the indicated time.

<p>Survival of <i>P. putida</i> (A, C) and <i>E. coli</i> (B, D) mutants defective in <i>alkA</i>, <i>alkB</i>, and <i>ada</i> genes treated with 20 mM MMS (A, B) or 5 µg/ml MNNG (C, D) for the indicated time.</p