The objectives of this work were to evaluate the genetic variability of Meloidogyne enterolobii by molecular markers, and develop species-specific molecular markers for application in detection. Sixteen M. enterolobii isolates from different geographical regions (Brazil and other countries) and hosts were used in this study. The identification and purification of the populations were carried out based on isoenzyme phenotype. The DNA amplification of the intergenic region (IGS) of the rDNA and of the region between the cytochrome oxidase subunit II (COII) and 16S rRNA genes (mtDNA) produced specific fragments of the expected size for this nematode, i.e. 780 and 705 bp, respectively. Intraspecific variability among the isolates was evaluated with three different neutral molecular markers: AFLP, ISSR and RAPD. The results showed a low level of diversity among the isolates tested, indicating that M. enterolobii is a genetically homogeneous root-knot nematode species. The RAPD method allowed the identification of a species-specific RAPD fragment for M. enterolobii. This fragment was cloned and sequenced, and from the sequence obtained, a set of primers was designed and tested. The amplification of a 520-bp-long fragment occurred only for the 16 isolates of M. enterolobii and not for the 10 other Meloidogyne species tested. In addition, positive detection was achieved in a single individual female, egg-mass and second stage juvenile of this nematode. This SCAR species-specific marker for M. enterolobii represents a new molecular tool to be used in the detection of this nematode from field samples and as a routine diagnostic test for quarantine devices.

Bacteriocins are ribosomally synthesized peptides that have bacteriostatic or bactericidal effects on other bacteria. The use of bacteriocins has emerged as an important strategy to increase food security and to minimize the incidence of foodborne diseases, due to its minimal impact on the nutritional and sensory properties of food products. Gramnegative bacteria are naturally resistant to the action of bacteriocins produced by Gram-positive bacteria, which are widely explored in foods. However, these microorganisms can be sensitized by mild treatments, such as the use of chelating agents, by treatment with plant essential oils or by physical treatments such as heating, freezing or high pressure processing. This sensitization is important in food microbiology, because most pathogens that cause foodborne diseases are Gram-negative bacteria. However, the effectiveness of these treatments is influenced by several factors, such as pH, temperature, the composition of the food and target microbiota. In this review, we comment on the main methods used for the sensitization of Gram-negative bacteria, especially Salmonella, to improve the action of bacteriocins produced by Gram-positive bacteria.

Meloidogyne incognita is one of the most polyphagous species of root-knot nematodes occurring in Brazil and worldwide. Eight M. incognita isolates were studied, representing two enzymatic phenotypes (esterase and malate desydrogenase: I1/N1, I2/N1) and four cryptic Meloidogyne sp.1 (S2/N1) isolates, representing one cytological type (3n040-46). Three M. hispanica isolates (Hi3/N1, 2n032-36) and two of an atypical Meloidogyne sp.2 (S2a/N3, 3n040-44) were included in this study for comparison. All isolates were tested with three M. incognita-specific molecular markers. The primer pairs B06F/R, miF/R and incK14F/R amplified three species-specific fragments of 1,200 bp, 955 bp and 399 bp, respectively for M. incognita and Meloidogyne sp.1 isolates. No amplification occurred in the M. hispanica and Meloidogyne sp.2 isolates, except with primers miF/R (1,650 bp). The genetic variability of the Meloidogyne spp. isolates was evaluated, using RAPD and ISSR markers. The phylogenetic analyses revealed two strongly supported monophyletic clades: clade I, consisting of M. hispanica and the atypical Meloidogyne sp.2 isolates, and clade II, clustering together all M. incognita and the Meloidogyne sp.1 isolates. Considering the biometrical, cytological and molecular approaches, it was possible to conclude that the isolates with three enzymatic phenotypes (I1/N1, I2/N1 and S2/N1) presented the characteristics described for M. incognita. Some correlations were detected between the isozymatic phenotypes and the tree topology (S2a/N3, Hi3/N1, I1/N1, S2/N1), but no strict correlation could be observed for the phenotype I2/N1 and one isolate of S2/N1. Morphologically, the Msp.2 isolates differ from M. incognita and M.

Meloidogyne paranaensis is one of the most destructive root-knot nematode (RKN) species parasitizing coffee in Brazil and in the Americas generally. The objectives of this study were to assess the genetic variability, aggressiveness and virulence of seven different M. paranaensis populations on susceptible and resistant Coffea spp. All seven RKN populations were identified by biochemical and molecular methods. Coffee seedlings were inoculated in the greenhouse, and the nematode reproduction factor was used to infer their reproduction on coffee genotypes. Phylogenetic studies showed a low genetic variability in M. paranaensis populations, regardless of the existence of three esterase phenotypes (Est P1, P2 and P2a), except for the population Est P2a from Guatemala, which is genetically different from other M. paranaensis populations from Brazil. The Est P2a and Est P2 (Herculândia, SP, Brazil) populations were the most aggressive on two susceptible C. arabica cultivars under greenhouse conditions. None of the M. paranaensis populations were virulent on resistant coffee genotypes, confirming their resistance to the seven M. paranaensis populations tested. The resistant coffee cultivars, namely Clone 14 INCAPER, Catua ı Vermelho 9 Amphillo MR2161 (E1 16-5 III), Apoatã IAC 2258, Timor Hybrid UFV 408-01 (E1 6-6 II) and IPR 100, exhibited segregation for resistance in the ratio of 0%, 2.4%, 12%, 26% and 29%, respectively. These are promising results, because they validate resistance against several M. paranaensis populations in different Coffea spp. genetic resources, which can be used in breeding programmes or as rootstocks, such as Apoatã IAC 2258 and Clone 14 INCAPER.