Industrial biotechnology is a continuously expanding field focused on the application of microorganisms to produce chemicals using renewable sources as substrates. Currently, an increasing interest in new versatile processes, able to utilize a variety of substrates to obtain diverse products, can be observed. A robust microbial strain is critical in the creation of such processes. Lactic acid bacteria (LAB) are used to produce a wide variety of chemicals with high commercial interest. Lactic acid (LA) is the most predominant industrial product obtained from LAB fermentations, and its production is forecasted to rise as the result of the increasing demand of polylactic acid. Hence, the creation of new ways to revalorize LA production processes is of high interest and could further enhance its economic value. Therefore, this review explores some co-products of LA fermentations, derived from LAB, with special focus on bacteriocins, lipoteichoic acid, and probiotics. Finally, a multi-product process involving LA and the other compounds of interest is proposed.Fermentation 2020, 6, 23 2 of 21 by-products, were proposed to improve industrial single-product biotechnological processes [2]. One example is the use of cellulose and hemicellulose from sugarcane bagasse (a residue obtained during the bioethanol production and usually used for energy generation) for the production of value-added chemicals [3]. Likewise, glycerol, a by-product of the biodiesel industry with low value in the market, is targeted as a molecule of interest in fermentation processes. Such multi-product processes, based on the conversion of renewable materials into biobased products, are known as biorefineries [4].The study, development, and application of robust microbial strains, able to utilize a variety of substrates and to produce a wide range of products, is considered a milestone in the development of biorefineries [4]. Lactic acid bacteria (LAB) are a diverse group with recognized potential for the development of integrated biorefineries [5]. LAB are non-sporulating, non-motile, acid-tolerant, non-respiring but aerotolerant, catalase-negative, Gram-positive cocci or rods. They are characterized by the production of lactic acid (LA) as the major end metabolic product of carbohydrate fermentation [6][7][8][9]. Given the lack of a functional respiratory system, LAB obtain energy through substrate-level phosphorylation following two metabolic pathways for hexose fermentation, i.e., homofermentative and heterofermentative. As shown in Figure 1, the first pathway is based on glycolysis with the production of mainly LA, whereas the second one, known as the pentose phosphate pathway, is characterized for the production of CO 2 and ethanol, or acetate in addition to LA [6].

The aim of this research was the genetic characterization of 218 accessions of Cucurbita moschata Duchesne, a squash, and its relationship with morphological characteristics of agronomic interest, which are part of the international collection conserved at Tropical Agricultural Research and Higher Education Center (CATIE), Costa Rica. The majority of the accessions came from Mexico and Central America; single genotypes from Curaao, Colombia, Peru and the Russian Federation were also included. The polymerase chain reaction (PCR) and single strand conformation polymorphism analysis (SSCP) were used for the analysis of the regions amplified with ITS1-ITS2 nuclear primers and tRNL-F chloroplast primers. Haplotypes were constructed according to band patterns in SSCP gels. Twenty-five haplotypes were found using the ITS1-ITS2 markers, and 24 haplotypes were found with the tRNL-F markers. Unique haplotypes were found with both markers. Two individuals of each tRNL-F haplotype were sequenced. The results indicated a high level of genetic diversity in CATIE squash collection. Using ITS1-ITS2 primers, it was found that the number of haplotypes was independent of the geographical source of the accession, and haplotypes were distributed randomly throughout the study area. Mexico had the highest values of total heterozygosity (HE), genetic diversity (H) and Shannon index (I) while Panama showed the lowest values. Sequences obtained from tRNL-F intergenic marker showed the highest diversity index values were present in the group of additional sequences and Mexico, and lower values were observed for Nicaragua, Guatemala and Panama. PCoA based on morphological data showed three groups and by ANOSIM (R) all group differences were significant. Results obtained in this study suggest that high diversity is a characteristic of C. moschata from Mesoamerica.

The tobacco whitefly Bemisia tabaci (Gennadius) cryptic species complex and of the greenhouse whitefly Trialeurodes vaporariorum (Westwood) are extensively reported as destructive pests in vegetable crops worldwide. A survey was conducted in 2011 and 2012 to determine the occurrence and genetic diversity present in the populations of these whiteflies in the major vegetable production areas of Costa Rica. Insect samples were collected from sweet pepper (Capsicum annuum L.), tomato (Solanum lycopersicum L.), common bean (Phaseolus vulgaris L.) and weeds present in commercial crops either in open field or greenhouse conditions. PCR‐RFLP analysis of mitochondrial cytochrome c oxidase subunit 1 gene (mtCOI) sequences of 621 whitefly individuals confirmed the presence of the Mediterranean (MED) type of the B. tabaci and of T. vaporariorum in most sampled regions. Also, individuals of the Middle East‐Asia Minor 1 (MEAM1) type of the B. tabaci were observed in low numbers. Contingency analyses based on type of crop, geographical region, whitefly species, year of collection and production system confirmed that T. vaporariorum was the most frequent species in vegetable production areas in Costa Rica, both in greenhouses and in open fields. B. tabaci MED is likely spreading to new areas of the country, whereas B. tabaci MEAM1 was mostly absent or rarely found. Comparisons of mtCOI sequences from B. tabaci individuals revealed the presence of four B. tabaci sequence haplotypes (named MED‐i, MED‐ii, MEAM1‐i, MEAM1‐xviii) in Costa Rica, three of them identical to B. tabaci haplotypes previously reported in the Western Hemisphere and other parts of the world. Analysis of sequences of T. vaporariorum individuals revealed a more complex population with the presence of 11 haplotypes, two of which were identical to T. vaporariorum sequences reported from other countries.

Begomoviruses (genus Begomovirus, family Geminiviridae) have emerged as important plant pathogens in tropical and subtropical regions worldwide. Although these viruses were reported during the 1970s in Costa Rica, they are still poorly known. Therefore, the objective of this study was to analyse the diversity and distribution of begomoviruses in commercial tomato and sweet pepper fields from different agricultural production systems of the major growing regions of Costa Rica. A total of 651 plants were randomly sampled from greenhouses and open field crops during 2011 and 2012 in three different geographical locations. The bipartite begomoviruses Tomato yellow mottle virus, Tomato leaf curl Sinaloa virus and Pepper golden mosaic virus, and the monopartite begomovirus Tomato yellow leaf curl virus were detected in the collected samples. The complete genome of isolates from each species was cloned and sequenced. The frequency of detection of these four begomoviruses in the analysed samples ranged from 0 to 9%, the presence, and the prevalent virus varied largely according to the geographical location, the host (tomato and pepper), and the production system (greenhouses or open fields). An association between geographical region and begomovirus species was observed suggesting that in Costa Rica the heterogeneity on climate, topography and agricultural system might influence the distribution of begomovirus species in the country. A broader survey needs to be conducted to confirm it, although these preliminary results may contribute to the management of begomoviruses in Costa Rica.

One of the most important invasive and harmful members of the genus Begomovirus (family Geminiviridae) is the monopartite Tomato yellow leaf curl virus (TYLCV), which is widespread over the world associated with tomato yellow leaf curl disease (TYLCD). Tomato (Solanum lycopersicum) plants infected with TYLCV show upward leaf curling and yellowing. In Latin America, isolates of TYLCV have been reported from Cuba, the Dominican Republic, Mexico and Puerto Rico (1), Guatemala (GenBank Accession No. GU355941), and Venezuela (partial genome sequence DQ302033). In Costa Rica, only isolates of the bipartite begomoviruses Tomato leaf curl Sinaloa virus (TLCSiV) (3) and Tomato yellow mottle virus (KC176780, KC176781) have been reported infecting tomatoes. During a survey conducted in 2012, similar begomovirus-like symptoms (leaf yellowing and upward leaf curling) were observed in tomato plants of five commercial growing areas in the Central Valley (Grecia region) of Costa Rica. In total, 65 tomato samples were randomly collected, 14 from greenhouses and 41 from open fields. Symptoms of upward leaf curling and yellowing were observed in three samples. Total DNA was extracted from collected samples and tested by dot blot hybridization using a probe to the coat protein (CP) gene of a Guatemalan isolate of Bean golden yellow mosaic virus (3). Only the three symptomatic samples tested positive, which represents an incidence of 14% (2 samples) in greenhouses and 2.4% (1 sample) in open field crops. These samples were subjected to rolling circle amplification (RCA) for viral circular genome amplification (2). The amplified products were then digested with MspI restriction endonuclease, which resulted into DNA fragments of 2,320 and 458 bp for all three samples. This suggested infection with a monopartite begomovirus. In order to obtain the full-length clone, the RCA product of two samples (5240 and 5241) was digested with BamHI, and the ~2.8 kb DNA fragment was cloned into pBluescript II SK(+) (Stratagene, La Jolla, CA) vector. After transformation of Escherichia coli DH5α, recombinant plasmids with inserts of expected size were selected and the insert was sequenced by primer walking (Macrogen Inc., Korea). The inserts of three clones from the two samples (CR:5240-16:2012, CR:5240-17:2012, and CR:5241-14:2012) were sequenced (deposited in GenBank as KF533855, KF533856, and KF533857, respectively). Sequences were all 2,781 nt long and shared 100% identity between themselves (1-nt mismatch between CR:5240-16:2012 and CR:5240-17:2012, and CR:5240-16:2012 and CR:5241-14:2012; and 2-nt mismatches between CR:5240-17:2012 and CR:5241-14:2012) and 99% with the sequence of Tomato yellow leaf curl virus-Israel[Japan:Haruno:2005] (TYLCV-IL[JR:Har:05]) (AB192966). These sequences represented full length genomes of isolates of the monopartite begomovirus TYLCV-IL and grouped in a phylogenetic clade (4) that comprised TYLCV-IL isolates reported from Asia (China and Japan) and from Mexico, while more distantly related to the clade comprising TYLCV-IL isolates reported from Central America (Cuba, Guatemala, Puerto Rico) and the United States, suggesting a distinct introduction event in Costa Rica. This is the first report of the presence of TYLCV in Costa Rica, therefore it is imperative to study the incidence and geographical spread of this virus in the country as well as its genetic diversity, since TYLCV infections might lead to significant yield losses, as reported in other countries. References: (1) A. M. Idris et al. Plant Dis. 83:303, 1999. (2) A. K. Inoue-Nagata et al. J. Virol. Methods 116:209, 2004. (3) M. K. Nakla et al. Acta Hortic. 695:277, 2005. (4) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011.