Negative Correlation between Individual-Insect-Level Virulence and Colony-Level Virulence of Paenibacillus larvae , the Etiological Agent of American Foulbrood of Honeybees

Abstract

Paenibacillus larvae is the etiological agent of American foulbrood (AFB) in honeybees. Recently, different genotypes of P. larvae (ERIC I to ERIC IV) were defined, and it was shown that these genotypes differ inter alia in their virulence on the larval level. On the colony level, bees mitigate AFB through the hygienic behavior of nurse bees. Therefore, we investigated how the hygienic behavior shapes P. larvae virulence on the colony level. Our results indicate that P. larvae virulence on the larval level and that on the colony level are negatively correlated.American foulbrood (AFB) is among the economically most important honeybee diseases. The etiological agent of AFB is the gram-positive, spore-forming bacterium Paenibacillus larvae (9). The extremely tenacious spores are the infectious form of this organism. These spores drive disease transmission within colonies (11), as well as between colonies as soon as they end up in the honey stores of an infected colony (12).The species P. larvae can be subdivided into four different genotypes designated ERIC I to ERIC IV based on results from repetitive-element PCR (20) using enterobacterial repetitive intergenic consensus (ERIC) primers (9, 10), with P. larvae ERIC I and ERIC II being the two practically most important genotypes (1, 2, 9, 10, 13, 16). The four genotypes were shown previously to differ in phenotype, including virulence on the larval level (8, 9). While larvae infected with genotypes ERIC II to ERIC IV were killed within only 6 to 7 days, it took P. larvae ERIC I around 12 to 14 days to kill all infected individuals. Therefore, genotype ERIC I was considered to be less virulent and the other three genotypes were considered to be highly virulent (7-9) on the larval level.P. larvae is an obligately killing pathogen which must kill its host to be transmitted. The virulence of such an obligate killer is thought to be determined primarily by two factors, (i) the probability of infecting a host and (ii) the time to host death (6). The problem of ensuring a high enough probability of infecting the next host is solved for P. larvae by (i) the tenacious exospores, which remain infectious for over half a century (17) and, therefore, can wait for decades for the next host to pass by, and (ii) a high pathogen reproduction rate (23) and, thus, the production of an extremely high number of spores within each infected larva.For evaluating the second factor determining P. larvae virulence, the time to host death, it is important to consider the two levels of honeybee hosts, the level of the individual larva dying from AFB and the level of the colony succumbing to AFB.The virulence of P. larvae genotypes on the larval level has been analyzed recently (8, 9). We have now determined the colony-level virulence for the two most common and practically important (10, 16) genotypes of P. larvae, ERIC I and ERIC II, significantly differing in virulence on the larval level (8). We will discuss how the time to larval death relates to the time to colony death and how the hygie...

Molecular pathogenesis of American Foulbrood: how Paenibacillus larvae kills honey bee larvae

“…However, it seems that the infected larva has no means to successfully combat infection and to escape being killed by P. larvae. Hence, individual immune responses though elicited by infection [46] are obviously not effective leaving it to the social immune response to deal with the infection at colony level [10,[47][48][49][50][51]. Breeding for hygienic behavior could, therefore, provide a solution to the AFB problem.…”

“…Strains of ERIC II-IV are fast killers: 100% of all infected larvae are already dead at day 6-7 post infection while ERIC I isolates need 10-12 days to kill 100% of all infected larvae. These differences in virulence at the individual larval level translate into differences at the colony level [10] which are relevant for the clinical diagnosis of AFB [11,12]. Recently, the ERIC-profile was additionally verified by state-of-theart analytical approaches like Multi Locus Sequence Typing (MLST) [13 ] and MALDI-ToF mass spectrometry [14].…”

Standard methods for American foulbrood research

“…The understanding of phenotypic differences within the species P. larvae (Genersch and Otten, 2003;Genersch et al, 2005;Neuendorf et al, 2004;Rauch et al, 2009) and the role of genotype-specific putative virulence factors (Fünfhaus et al, 2009;Fünfhaus and Genersch, 2012) have been hampered by the lack of molecular tools allowing genetic manipulation of this pathogen.…”

Expression of larval jelly antimicrobial peptide defensin1 in Apis mellifera colonies

“…Less than 10 spores in larval diet may suffice to infect midgut of 12-36 hour old larvae and cause their death and fatal infection in colony due to massive spore production (one decomposed larva produces 2 × 10 9 spores) and their easy transmission within colony (Bailey & Ball 1991;Shimanuki et al 1992;Hansen & Brødsgaard 1999;Fries & Camazine 2001;Lindström et al 2008). Incidence, progression and severity of AFB in colonies are affected by virulence of P. larvae, bee defense abilities, environmental conditions and stress (Genersch et al 2005;Rauch et al 2009;Genersch 2010). Colonies exhibit different degrees of resistance against AFB (Rothenbuhler & Thompson 1956;Bambrick 1964;Hansen & Brødsgaard 1999) which might be associated with the efficacy of individual and social/collective defense traits of larvae and adult bees (Cremer et al 2007;Wilson-Rich et al 2009;Evans & Spivak 2010).…”