Авиламицин инструкция по применению в ветеринарии

2.3.1 Lactobacillus sp. probiotics

Following the Metchnikoff studies of Lactobacilli and their use as probiotics, Lactobacilliare are now the most commonly used probiotic bacterial strains. Several experiments have shown significant effects of Lactobacillus probiotic on weight in animals and humans, and these effects vary according to the species [25,26]. Some species were linked to weight gain, whereas others species were associated with weight loss. Meta-analyses revealed that Lactobacillus acidophilus, Lactobacillus fermentum and Lactobacillus ingluviei administration resulted in significant weight gain whereas Lactobacillus plantarum and Lactobacillus gasseri are associated with weight loss [26]. To date, L. acidophilus, L. fermentum, L. johnsonni, L. paracasei, L. plantarum, L. reuteri, L. rhamnosus and L. salivarius are the most commercialized Lactobacilli [27]. Concentrates of these bacteria are usually freeze-dried, spray-dried, or micro-encapsulated and are typically incorporated into dairy products in humans or in water in animals. Lactobacillus probiotics have been successfully used for the treatment of diarrhea, for irritable bowel syndrome, inflammatory bowel disease, and pouchitis [28]. Many commercial Lactobacilli probiotics products are currently available on the market worldwide and are rapidly gaining in popularity [29,30].

2.3.2 Poultry

2.3.3 Ruminants

The most commonly marketed products for ruminants are live yeast (Saccharomyces cerevisiae) preparations. In dairy ruminants, live yeasts have been shown to improve performance, the most consistent effects being an increase in dry matter intake and milk production. Ruminal acidosis is a common digestive disorder in high-producing dairy or beef cattle and is responsible for a decrease in animal performance. In vitro studies have reported that live yeasts could influence the balance of lactate-metabolizing bacteria, by limiting lactate production by Streptococcus bovis and favoring lactate uptake by Megasphaeraelsdenii or Selenomonasruminantium [39]. Moreover, S. cerevisiae could prevent pH decrease by stimulating certain populations of ciliate protozoa [40]. Feed efficiency can also be significantly improved after probiotic feeding [41]. The carcass-based gain/feed ratios also tended to be better for animals receiving the probiotic treatment [42]. Lactobacillus sp. treatment also resulted in higher metabolic activity, lower levels of non-esterified fatty acids, triglycerides, urea, and an increase in alkaline phosphatase and creatine kinase levels [43].

2.3.4 Piglets

2.3.5 Aquaculture

A wide variety of Gram-negative bacteria play a role as putative probiotics in aquaculture [49]. Probiotic administrations have been widely applied via water routine or feed additives with either single or a combination of probiotics or even a mixture with prebiotics or other immunostimulants. Probiotic administration varies from direct oral/water routine or feed additives, in which the former is considered the most practical method for prawn probiotics. Normally, probiotics can be added directly into culture water as water additives, bathed in bacterial suspension [50]. Endospore-forming members of Bacillus genera including Bacillus subtilis are commonly used in aquaculture [50]. Moreover, several yeasts have been proven to provide benefits to aquatic animals. Saccharomyces cerevisiae has been recognized to have potential as a substitute for live feed production of fish whereas the marine yeast Yarrowialipolytica has improved the survival and growth of pearl oysters [50]. However, appropriate probiotic levels depend on the probiotic species, fish species and their physiological status, rearing conditions and specific goal of the applications.

16S rRNA Methyltransferases

Intriguingly, the 16S rRNA methyltransferases identified in aminoglycoside producers show low sequence identity with those circulating in pathogenic strains [91]. Aminoglycoside producers possess two families: the Kgm family (KgmB, Sgm, GrmA, etc.) and the Kam family (KamA, KamB, and KamC), which methylate G1405 and A1408, respectively. By contrast, pathogenic bacteria have acquired two families independent of those from the aminoglycoside producers. The Arm family of methyltransferases comprises six members (ArmA and RmtA through RmtE), which methylate G1405 and thus produce an aminoglycoside resistance phenotype like that of the Kgm family [92,93]. The second family, the Pam family, currently comprises solely NpmA, which methylates A1408 like Kam methyltransferases [94]. Regardless of the family, methylation at either nucleotide drastically hinders the therapeutic use of aminoglycosides.

23S rRNA Methyltransferases

Unlike 16S rRNA methyltransferases, the 23S rRNA methyltransferases are more mechanistically diverse, comprising N-methyltransferases, radical SAM methyltransferases, and 2′-O-ribose methyltransferases. Except for the radical SAM methyltransferases, these enzymes are likely to facilitate an S_N2 mechanism for methylation of their nucleophilic targets [95,96]. Methylation of the 23S rRNA affects resistance against ten different antibiotic classes depending on the location and combination of methylation sites (Table 1). In many cases, a single methylation can cause resistance to multiple antibiotic classes due to the extensive overlap in their binding sites.

N-Methylations are the most numerous of the 23S rRNA modifications. One of the most well-known families is the Erm N-methyltransferases, which methylate the N6 position of A2058 at the mouth of the peptide exit tunnel [97]. Some Erm methyltransferases, such as ErmN monomethylate A2058, lead to robust lincosamide resistance and low-to-moderate resistance to macrolide and streptogramin B antibiotics, although they offer no resistance to the subset of macrolides known as the ketolides [98]. Others, such as ErmC dimethylate A2058, which creates strong resistance to all three classes (the ‘MLS_B’ phenotype), including the ketolides [84]. Intriguingly, ErmN also confers strong resistance to the macrolide tylosin when found in conjunction with the N-methyltransferase RlmA^II, which methylates G748 [99]. Finally, resistance to the orthosomycins avilamycin and evernimicin occurs through methylation of G2470 and G2535 by EmtA and AviRa, respectively. However, the latter is only effective if 2′-O-ribose methylation of U2497 by AviRb is also present [100].

Although less prevalent in the clinic, 2′-O-ribose methylation also plays a role in resistance to certain antibiotics (Figure 4A and Table 1). Two such cases exist, both used as intrinsic resistance mechanisms in the producer of the affected antibiotic: AviRb and Tsr. As noted above, AviRb (in conjunction with AviRa) is crucial for orthosomycin resistance. By contrast, Tsr protects the thiostrepton producer Streptomyces azureus from self-intoxication via ribose methylation of A1067, a nucleotide in the ribosome’s GTPase center [85,101]. While neither thiostrepton nor the orthosomycins are currently used in human medicine, their resistance mechanisms may affect any future antibiotic that targets similar binding sites.

Unlike the N-methyltransferases and 2′-O-methyltransferases, the radical SAM methyltransferases do not utilize an S_N2 mechanism to methylate their targets. Instead, they require the generation of a free radical to methylate relatively inert carbons. To date, only a single radical SAM methyltransferase, Cfr, is associated with antibiotic resistance [102]. Like other radical SAM methyltransferases, Cfr uses two SAM molecules to perform the methylation: the first to methylate a conserved cysteine in the active site and the second as a source for a 5′-deoxyadenosine radical. The radical is passed from 5′-deoxyadenosine to the methylated cysteine, which subsequently attacks the target nucleotide, ultimately transferring the methyl group [103]. Cfr methylates the C8 position of A2503 in the peptidyl transferase center, resulting in resistance to phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptogramin A (the ‘PhLOPS_A’ phenotype) [83]. Alarmingly, Cfr is readily mobile within pathogens and has recently been found in the same operon as the ErmB methyltransferase [104,105]; such a combination renders the ribosome resistant to all clinically relevant 50S ribosome-targeting antibiotics.

Resistance through the Loss of Methylation

In some instances, antibiotic resistance can occur via loss of endogenous methylation. The tuberactinomycin antibiotics capreomycin and viomycin bind at the interface of the 30S and 50S subunits in the intact ribosome, and ribose methylation of 16S rRNA C1409 and 23S rRNA C1920 is required for their antimycobacterial activity [106]. Inactivating mutations in the tlyA gene result in the loss of both methylations and the generation of capreomycin resistance in Mycobacterium tuberculosis [107]. In another example, inactivation of KsgA abolishes methylation of A1518 and A1519 in the 16S rRNA, leading to low-level kasugamycin resistance. Intriguingly, KsgA inactivation also dramatically accelerates the emergence of secondary high-level kasugamycin resistance [108]. Resistance via loss of methylation may prove to be challenging to overcome as it is the loss of an enzyme rather than the acquisition of a targetable resistance enzyme.

Overcoming Methylation-Dependent Resistance

Since methylation typically causes resistance by steric clashes with the antibiotic, one strategy to overcome this mechanism is by designing antibiotics that avoid the methylated nucleotide. The recently approved oxazolidinone tedizolid was designed with a shorter C5 substituent on its oxazolidinone ring relative to its predecessor linezolid (Figure 4B); this modification accommodates the methylation of A2503 and thus retains activity against cfr⁺ pathogens [109]. Another example is the ketolide telithromycin, which extends deeper into the peptide exit tunnel than the macrolides, allowing more extensive interaction with the 23S rRNA [110]. Increased interactions with the rRNA enable telithromycin to overcome resistance induced by monomethylation of A2058 [111]. Unfortunately, telithromycin is unable to overcome resistance caused by dimethylation of A2058.

In the cases of a single methylation causing multidrug resistance, it may be more pragmatic to target the methyltransferase itself, thus salvaging multiple antibiotic classes by inhibiting a single enzyme. ErmC has gained considerable attention in this regard due to the robust MLS_B phenotype [112–114]. Clancy et al. performed a traditional high-throughput screen of synthetic compounds against purified ErmC. Their most potent hit, UK-105730, has an IC₅₀ of 0.45 μM and demonstrated superior selectivity against other methyltransferases (IC₅₀ > 1 mM) (Figure 4C). Hajduk et al. also utilized a synthetic screen approach, but instead screened small-molecule fragments for enzyme binding by NMR followed by an extensive medicinal chemistry regimen to build on the hits. Their efforts yielded a 2-amino-4-(aminoindanyl)-1,3,5-triazine scaffold, the best of which showed a K_i of 4 μM and 10 μM against ErmAM and ErmC, respectively (Figure 4C). Finally, Foik et al. performed an in silico screen followed by in vitro experimentation to identify inhibitors that would bind both the SAM-binding pocket and the rRNA-binding pocket. Intriguingly, they identified a family of three related compounds that possessed different modes of inhibition (competitive, noncompetitive, and uncompetitive) (Figure 4C). Unfortunately, to date none of the identified Erm inhibitors have demonstrated rescue of macrolide activity in a mouse model of infection when tested.